Isolated human enzyme proteins, nucleic acid molecules encoding human enzyme proteins, and uses thereof

ABSTRACT

The present invention provides amino acid sequences of peptides that are encoded by genes within the human genome, the enzyme peptides of the present invention. The present invention specifically provides isolated peptide and nucleic acid molecules, methods of identifying orthologs and paralogs of the enzyme peptides, and methods of identifying modulators of the enzyme peptides.

FIELD OF THE INVENTION

[0001] The present invention is in the field of enzyme proteins that are related to the lanosterol synthase enzyme subfamily, recombinant DNA molecules, and protein production. The present invention specifically provides novel peptides and proteins that effect protein phosphorylation and nucleic acid molecules encoding such peptide and protein molecules, all of which are useful in the development of human therapeutics and diagnostic compositions and methods.

BACKGROUND OF THE INVENTION

[0002] Many human enzymes serve as targets for the action of pharmaceutically active compounds. Several classes of human enzymes that serve as such targets include helicase, steroid esterase and sulfatase, convertase, synthase, dehydrogenase, monoxygenase, transferase, kinase, glutanase, decarboxylase, isomerase and reductase. It is therefore important in developing new pharmaceutical compounds to identify target enzyme proteins that can be put into high-throughput screening formats. The present invention advances the state of the art by providing novel human drug target enzymes related to the lanosterol synthase subfamily.

[0003] Lanosterol Synthase

[0004] The novel human protein, and encoding gene, provided by the present invention is related to lanosterol synthase enzymes. Specifically, the protein provided by the present invention is 11 amino acids shorter in exon 4 compared with the art-known lanosterol synthase protein provided in Genbank gi4505027 (see the amino acid sequence alignment of the protein of the present invention against gi4505027 provided in FIG. 2).

[0005] Lanosterol synthase enzymes are important for catalyzing the cyclization of squalene-2,3-epoxide lanosterol, which is the parental compound of all mammalian steroids (Young et al., Human Genet 1996 May; 97(5):620-4). Baker et al. (Biochem Biophys Res Commun 1995 Aug. 4;213(1):154-60) cloned and characterized the human lanosterol synthase gene and found that it encoded a predicted 83 kDa protein of 732 amino acids; this amino acid sequence shared 36-40% identity with yeast and plant homologues and 83% identity with Rattus norvegicus lanosterol synthase. For a further review of the lanosterol synthase gene/protein, see Sung et al., Biol Pharm Bull 1995 October; 18(10): 1459-61.

[0006] Enzyme proteins, particularly members of the lanosterol synthase enzyme subfamily, are a major target for drug action and development. Accordingly, it is valuable to the field of pharmaceutical development to identify and characterize previously unknown members of this subfamily of enzyme proteins. The present invention advances the state of the art by providing previously unidentified human enzyme proteins, and the polynucleotides encoding them, that have homology to members of the lanosterol synthase enzyme subfamily. These novel compositions are useful in the diagnosis, prevention and treatment of biological processes associated with human diseases.

SUMMARY OF THE INVENTION

[0007] The present invention is based in part on the identification of amino acid sequences of human enzyme peptides and proteins that are related to the lanosterol synthase enzyme subfamily, as well as allelic variants and other mammalian orthologs thereof. These unique peptide sequences, and nucleic acid sequences that encode these peptides, can be used as models for the development of human therapeutic targets, aid in the identification of therapeutic proteins, and serve as targets for the development of human therapeutic agents that modulate enzyme activity in cells and tissues that express the enzyme. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus.

DESCRIPTION OF THE FIGURE SHEETS

[0008]FIG. 1 provides the nucleotide sequence of a cDNA molecule that encodes the enzyme protein of the present invention. (SEQ ID NO:1) In addition, structure and functional information is provided, such as ATG start, stop and tissue distribution, where available, that allows one to readily determine specific uses of inventions based on this molecular sequence. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus.

[0009]FIG. 2 provides the predicted amino acid sequence of the enzyme of the present invention. (SEQ ID NO:2) In addition structure and functional information such as protein family, function, and modification sites is provided where available, allowing one to readily determine specific uses of inventions based on this molecular sequence.

[0010]FIG. 3 provides genomic sequences that span the gene encoding the enzyme protein of the present invention. (SEQ ID NO:3) In addition structure and functional information, such as intron/exon structure, promoter location, etc., is provided where available, allowing one to readily determine specific uses of inventions based on this molecular sequence. As illustrated in FIG. 3, SNPs were identified at 56 different nucleotide positions.

DETAILED DESCRIPTION OF THE INVENTION

[0011] General Description

[0012] The present invention is based on the sequencing of the human genome. During the sequencing and assembly of the human genome, analysis of the sequence information revealed previously unidentified fragments of the human genome that encode peptides that share structural and/or sequence homology to protein/peptide/domains identified and characterized within the art as being a enzyme protein or part of a enzyme protein and are related to the lanosterol synthase enzyme subfamily. Utilizing these sequences, additional genomic sequences were assembled and transcript and/or cDNA sequences were isolated and characterized. Based on this analysis, the present invention provides amino acid sequences of human enzyme peptides and proteins that are related to the lanosterol synthase enzyme subfamily, nucleic acid sequences in the form of transcript sequences, cDNA sequences and/or genomic sequences that encode these enzyme peptides and proteins, nucleic acid variation (allelic information), tissue distribution of expression, and information about the closest art known protein/peptide/domain that has structural or sequence homology to the enzyme of the present invention.

[0013] In addition to being previously unknown, the peptides that are provided in the present invention are selected based on their ability to be used for the development of commercially important products and services. Specifically, the present peptides are selected based on homology and/or structural relatedness to known enzyme proteins of the lanosterol synthase enzyme subfamily and the expression pattern observed. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. The art has clearly established the commercial importance of members of this family of proteins and proteins that have expression patterns similar to that of the present gene. Some of the more specific features of the peptides of the present invention, and the uses thereof, are described herein, particularly in the Background of the Invention and in the annotation provided in the Figures, and/or are known within the art for each of the known lanosterol synthase family or subfamily of enzyme proteins.

[0014] Specific Embodiments

[0015] Peptide Molecules

[0016] The present invention provides nucleic acid sequences that encode protein molecules that have been identified as being members of the enzyme family of proteins and are related to the lanosterol synthase enzyme subfamily (protein sequences are provided in FIG. 2, transcript/cDNA sequences are provided in FIG. 1 and genomic sequences are provided in FIG. 3). The peptide sequences provided in FIG. 2, as well as the obvious variants described herein, particularly allelic variants as identified herein and using the information in FIG. 3, will be referred herein as the enzyme peptides of the present invention, enzyme peptides, or peptides/proteins of the present invention.

[0017] The present invention provides isolated peptide and protein molecules that consist of, consist essentially of, or comprise the amino acid sequences of the enzyme peptides disclosed in the FIG. 2, (encoded by the nucleic acid molecule shown in FIG. 1, transcript/cDNA or FIG. 3, genomic sequence), as well as all obvious variants of these peptides that are within the art to make and use. Some of these variants are described in detail below.

[0018] As used herein, a peptide is said to be “isolated” or “purified” when it is substantially free of cellular material or free of chemical precursors or other chemicals. The peptides of the present invention can be purified to homogeneity or other degrees of purity. The level of purification will be based on the intended use. The critical feature is that the preparation allows for the desired function of the peptide, even if in the presence of considerable amounts of other components (the features of an isolated nucleic acid molecule is discussed below).

[0019] In some uses, “substantially free of cellular material” includes preparations of the peptide having less than about 30% (by dry weight) other proteins (i.e., contaminating protein), less than about 20% other proteins, less than about 10% other proteins, or less than about 5% other proteins. When the peptide is recombinantly produced, it can also be substantially free of culture medium, i.e., culture medium represents less than about 20% of the volume of the protein preparation.

[0020] The language “substantially free of chemical precursors or other chemicals” includes preparations of the peptide in which it is separated from chemical precursors or other chemicals that are involved in its synthesis. In one embodiment, the language “substantially free of chemical precursors or other chemicals” includes preparations of the enzyme peptide having less than about 30% (by dry weight) chemical precursors or other chemicals, less than about 20% chemical precursors or other chemicals, less than about 10% chemical precursors or other chemicals, or less than about 5% chemical precursors or other chemicals.

[0021] The isolated enzyme peptide can be purified from cells that naturally express it, purified from cells that have been altered to express it (recombinant), or synthesized using known protein synthesis methods. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. For example, a nucleic acid molecule encoding the enzyme peptide is cloned into an expression vector, the expression vector introduced into a host cell and the protein expressed in the host cell. The protein can then be isolated from the cells by an appropriate purification scheme using standard protein purification techniques. Many of these techniques are described in detail below.

[0022] Accordingly, the present invention provides proteins that consist of the amino acid sequences provided in FIG. 2 (SEQ ID NO:2), for example, proteins encoded by the transcript/cDNA nucleic acid sequences shown in FIG. 1 (SEQ ID NO:1) and the genomic sequences provided in FIG. 3 (SEQ ID NO:3). The amino acid sequence of such a protein is provided in FIG. 2. A protein consists of an amino acid sequence when the amino acid sequence is the final amino acid sequence of the protein.

[0023] The present invention further provides proteins that consist essentially of the amino acid sequences provided in FIG. 2 (SEQ ID NO:2), for example, proteins encoded by the transcript/cDNA nucleic acid sequences shown in FIG. 1 (SEQ ID NO:1) and the genomic sequences provided in FIG. 3 (SEQ ID NO:3). A protein consists essentially of an amino acid sequence when such an amino acid sequence is present with only a few additional amino acid residues, for example from about 1 to about 100 or so additional residues, typically from 1 to about 20 additional residues in the final protein.

[0024] The present invention further provides proteins that comprise the amino acid sequences provided in FIG. 2 (SEQ ID NO:2), for example, proteins encoded by the transcript/cDNA nucleic acid sequences shown in FIG. 1 (SEQ ID NO:1) and the genomic sequences provided in FIG. 3 (SEQ ID NO:3). A protein comprises an amino acid sequence when the amino acid sequence is at least part of the final amino acid sequence of the protein. In such a fashion, the protein can be only the peptide or have additional amino acid molecules, such as amino acid residues (contiguous encoded sequence) that are naturally associated with it or heterologous amino acid residues/peptide sequences. Such a protein can have a few additional amino acid residues or can comprise several hundred or more additional amino acids. The preferred classes of proteins that are comprised of the enzyme peptides of the present invention are the naturally occurring mature proteins. A brief description of how various types of these proteins can be made/isolated is provided below.

[0025] The enzyme peptides of the present invention can be attached to heterologous sequences to form chimeric or fusion proteins. Such chimeric and fusion proteins comprise a enzyme peptide operatively linked to a heterologous protein having an amino acid sequence not substantially homologous to the enzyme peptide. “Operatively linked” indicates that the enzyme peptide and the heterologous protein are fused in-frame. The heterologous protein can be fused to the N-terminus or C-terminus of the enzyme peptide.

[0026] In some uses, the fusion protein does not affect the activity of the enzyme peptide per se. For example, the fusion protein can include, but is not limited to, enzymatic fusion proteins, for example beta-galactosidase fusions, yeast two-hybrid GAL fusions, poly-His fusions, MYC-tagged, HI-tagged and Ig fusions. Such fusion proteins, particularly poly-His fusions, can facilitate the purification of recombinant enzyme peptide. In certain host cells (e.g., mammalian host cells), expression and/or secretion of a protein can be increased by using a heterologous signal sequence.

[0027] A chimeric or fusion protein can be produced by standard recombinant DNA techniques. For example, DNA fragments coding for the different protein sequences are ligated together in-frame in accordance with conventional techniques. In another embodiment, the fusion gene can be synthesized by conventional techniques including automated DNA synthesizers. Alternatively, PCR amplification of gene fragments can be carried out using anchor primers which give rise to complementary overhangs between two consecutive gene fragments which can subsequently be annealed and re-amplified to generate a chimeric gene sequence (see Ausubel et al., Current Protocols in Molecular Biology, 1992). Moreover, many expression vectors are commercially available that already encode a fusion moiety (e.g., a GST protein). A enzyme peptide-encoding nucleic acid can be cloned into such an expression vector such that the fusion moiety is linked in-frame to the enzyme peptide.

[0028] As mentioned above, the present invention also provides and enables obvious variants of the amino acid sequence of the proteins of the present invention, such as naturally occurring mature forms of the peptide, allelic/sequence variants of the peptides, non-naturally occurring recombinantly derived variants of the peptides, and orthologs and paralogs of the peptides. Such variants can readily be generated using art-known techniques in the fields of recombinant nucleic acid technology and protein biochemistry. It is understood, however, that variants exclude any amino acid sequences disclosed prior to the invention.

[0029] Such variants can readily be identified/made using molecular techniques and the sequence information disclosed herein. Further, such variants can readily be distinguished from other peptides based on sequence and/or structural homology to the enzyme peptides of the present invention. The degree of homology/identity present will be based primarily on whether the peptide is a functional variant or non-functional variant, the amount of divergence present in the paralog family and the evolutionary distance between the orthologs.

[0030] To determine the percent identity of two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment and non-homologous sequences can be disregarded for comparison purposes). In a preferred embodiment, at least 30%, 40%, 50%, 60%, 70%, 80%, or 90% or more of the length of a reference sequence is aligned for comparison purposes. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position (as used herein amino acid or nucleic acid “identity” is equivalent to amino acid or nucleic acid “homology”). The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences.

[0031] The comparison of sequences and determination of percent identity and similarity between two sequences can be accomplished using a mathematical algorithm. (Computational Molecular Biology, Lesk, A. M., ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, D. W., ed., Academic Press, New York, 1993; Computer Analysis of Sequence Data, Part 1, Griffin, A. M., and Griffin, H. G., eds., Humana Press, New Jersey, 1994; Sequence Analysis in Molecular Biology, von Heinje, G., Academic Press, 1987; and Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991). In a preferred embodiment, the percent identity between two amino acid sequences is determined using the Needleman and Wunsch (J. Mol. Biol. (48):444-453 (1970)) algorithm which has been incorporated into the GAP program in the GCG software package (available at http://Hwww.gcg.com), using either a Blossom 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6. In yet another preferred embodiment, the percent identity between two nucleotide sequences is determined using the GAP program in the GCG software package (Devereux, J., et al., Nucleic Acids Res. 12(1):387 (1984)) (available at http://www.gcg.com), using a NWSgapdna.CMP matrix and a gap weight of 40, 50, 60, 70, or 80 and a length weight of 1, 2, 3, 4, 5, or 6. In another embodiment, the percent identity between two amino acid or nucleotide sequences is determined using the algorithm of E. Myers and W. Miller (CABIOS, 4:11-17 (1989)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4.

[0032] The nucleic acid and protein sequences of the present invention can further be used as a “query sequence” to perform a search against sequence databases to, for example, identify other family members or related sequences. Such searches can be performed using the NBLAST and XBLAST programs (version 2.0) of Altschul, et al. (J. Mol. Biol. 215:403-10 (1990)). BLAST nucleotide searches can be performed with the NBLAST program, score=100, wordlength=12 to obtain nucleotide sequences homologous to the nucleic acid molecules of the invention. BLAST protein searches can be performed with the XBLAST program, score=50, wordlength=3 to obtain amino acid sequences homologous to the proteins of the invention. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al. (Nucleic Acids Res. 25(17):3389-3402 (1997)). When utilizing BLAST and gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used.

[0033] Full-length pre-processed forms, as well as mature processed forms, of proteins that comprise one of the peptides of the present invention can readily be identified as having complete sequence identity to one of the enzyme peptides of the present invention as well as being encoded by the same genetic locus as the enzyme peptide provided herein. The gene encoding the novel enzyme of the present invention is located on a genome component that has been mapped to human chromosome 21 (as indicated in FIG. 3), which is supported by multiple lines of evidence, such as STS and BAC map data.

[0034] Allelic variants of a enzyme peptide can readily be identified as being a human protein having a high degree (significant) of sequence homology/identity to at least a portion of the enzyme peptide as well as being encoded by the same genetic locus as the enzyme peptide provided herein. Genetic locus can readily be determined based on the genomic information provided in FIG. 3, such as the genomic sequence mapped to the reference human. The gene encoding the novel enzyme of the present invention is located on a genome component that has been mapped to human chromosome 21 (as indicated in FIG. 3), which is supported by multiple lines of evidence, such as STS and BAC map data. As used herein, two proteins (or a region of the proteins) have significant homology when the amino acid sequences are typically at least about 70-80%, 80-90%, and more typically at least about 90-95% or more homologous. A significantly homologous amino acid sequence, according to the present invention, will be encoded by a nucleic acid sequence that will hybridize to a enzyme peptide encoding nucleic acid molecule under stringent conditions as more fully described below.

[0035]FIG. 3 provides information on SNPs that have been found in the gene encoding the enzyme of the present invention. SNPs were identified at 56 different nucleotide positions, including a non-synonymous coding SNP at position 36001 (protein position 631). Changes in the amino acid sequence caused by these SNPs is indicated in FIG. 3 and can readily be determined using the universal genetic code and the protein sequence provided in FIG. 2 as a reference. Some of these SNPs, located outside the ORF and in introns, may affect gene transcription.

[0036] Paralogs of a enzyme peptide can readily be identified as having some degree of significant sequence homology/identity to at least a portion of the enzyme peptide, as being encoded by a gene from humans, and as having similar activity or function. Two proteins will typically be considered paralogs when the amino acid sequences are typically at least about 60% or greater, and more typically at least about 70% or greater homology through a given region or domain. Such paralogs will be encoded by a nucleic acid sequence that will hybridize to a enzyme peptide encoding nucleic acid molecule under moderate to stringent conditions as more fully described below.

[0037] Orthologs of a enzyme peptide can readily be identified as having some degree of significant sequence homology/identity to at least a portion of the enzyme peptide as well as being encoded by a gene from another organism. Preferred orthologs will be isolated from mammals, preferably primates, for the development of human therapeutic targets and agents. Such orthologs will be encoded by a nucleic acid sequence that will hybridize to a enzyme peptide encoding nucleic acid molecule under moderate to stringent conditions, as more fully described below, depending on the degree of relatedness of the two organisms yielding the proteins.

[0038] Non-naturally occurring variants of the enzyme peptides of the present invention can readily be generated using recombinant techniques. Such variants include, but are not limited to deletions, additions and substitutions in the amino acid sequence of the enzyme peptide. For example, one class of substitutions are conserved amino acid substitution. Such substitutions are those that substitute a given amino acid in a enzyme peptide by another amino acid of like characteristics. Typically seen as conservative substitutions are the replacements, one for another, among the aliphatic amino acids Ala, Val, Leu, and lie; interchange of the hydroxyl residues Ser and Thr; exchange of the acidic residues Asp and Glu; substitution between the amide residues Asn and Gln; exchange of the basic residues Lys and Arg; and replacements among the aromatic residues Phe and Tyr. Guidance concerning which amino acid changes are likely to be phenotypically silent are found in Bowie et al., Science 247:1306-1310 (1990).

[0039] Variant enzyme peptides can be fully functional or can lack function in one or more activities, e.g. ability to bind substrate, ability to phosphorylate substrate, ability to mediate signaling, etc. Fully functional variants typically contain only conservative variation or variation in non-critical residues or in non-critical regions. FIG. 2 provides the result of protein analysis and can be used to identify critical domains/regions. Functional variants can also contain substitution of similar amino acids that result in no change or an insignificant change in function. Alternatively, such substitutions may positively or negatively affect function to some degree.

[0040] Non-functional variants typically contain one or more non-conservative amino acid substitutions, deletions, insertions, inversions, or truncation or a substitution, insertion, inversion, or deletion in a critical residue or critical region.

[0041] Amino acids that are essential for function can be identified by methods known in the art, such as site-directed mutagenesis or alanine-scanning mutagenesis (Cunningham et al., Science 244:1081-1085 (1989)), particularly using the results provided in FIG. 2. The latter procedure introduces single alanine mutations at every residue in the molecule. The resulting mutant molecules are then tested for biological activity such as enzyme activity or in assays such as an in vitro proliferative activity. Sites that are critical for binding partner/substrate binding can also be determined by structural analysis such as crystallization, nuclear magnetic resonance or photoaffinity labeling (Smith et al., J. Mol. Biol. 224:899-904 (1992); de Vos et al. Science 255:306-312 (1992)).

[0042] The present invention further provides fragments of the enzyme peptides, in addition to proteins and peptides that comprise and consist of such fragments, particularly those comprising the residues identified in FIG. 2. The fragments to which the invention pertains, however, are not to be construed as encompassing fragments that may be disclosed publicly prior to the present invention.

[0043] As used herein, a fragment comprises at least 8, 10, 12, 14, 16, or more contiguous amino acid residues from a enzyme peptide. Such fragments can be chosen based on the ability to retain one or more of the biological activities of the enzyme peptide or could be chosen for the ability to perform a function, e.g. bind a substrate or act as an immunogen. Particularly important fragments are biologically active fragments, peptides that are, for example, about 8 or more amino acids in length. Such fragments will typically comprise a domain or motif of the enzyme peptide, e.g., active site, a transmembrane domain or a substrate-binding domain. Further, possible fragments include, but are not limited to, domain or motif containing fragments, soluble peptide fragments, and fragments containing immunogenic structures. Predicted domains and functional sites are readily identifiable by computer programs well known and readily available to those of skill in the art (e.g., PROSITE analysis). The results of one such analysis are provided in FIG. 2.

[0044] Polypeptides often contain amino acids other than the 20 amino acids commonly referred to as the 20 naturally occurring amino acids. Further, many amino acids, including the terminal amino acids, may be modified by natural processes, such as processing and other post-translational modifications, or by chemical modification techniques well known in the art. Common modifications that occur naturally in enzyme peptides are described in basic texts, detailed monographs, and the research literature, and they are well known to those of skill in the art (some of these features are identified in FIG. 2).

[0045] Known modifications include, but are not limited to, acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent crosslinks, formation of cystine, formation of pyroglutamate, formylation, gamma carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination.

[0046] Such modifications are well known to those of skill in the art and have been described in great detail in the scientific literature. Several particularly common modifications, glycosylation, lipid attachment, sulfation, gamma-carboxylation of glutamic acid residues, hydroxylation and ADP-ribosylation, for instance, are described in most basic texts, such as Proteins—Structure and Molecular Properties, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York (1993). Many detailed reviews are available on this subject, such as by Wold, F., Posttranslational Covalent Modification of Proteins, B. C. Johnson, Ed., Academic Press, New York 1-12 (1983); Seifter et al. (Meth. Enzymol. 182: 626-646 (1990)) and Rattan et al. (Ann. N.Y. Acad. Sci. 663:48-62 (1992)).

[0047] Accordingly, the enzyme peptides of the present invention also encompass derivatives or analogs in which a substituted amino acid residue is not one encoded by the genetic code, in which a substituent group is included, in which the mature enzyme peptide is fused with another compound, such as a compound to increase the half-life of the enzyme peptide (for example, polyethylene glycol), or in which the additional amino acids are fused to the mature enzyme peptide, such as a leader or secretory sequence or a sequence for purification of the mature enzyme peptide or a pro-protein sequence.

[0048] Protein/Peptide Uses

[0049] The proteins of the present invention can be used in substantial and specific assays related to the functional information provided in the Figures; to raise antibodies or to elicit another immune response; as a reagent (including the labeled reagent) in assays designed to quantitatively determine levels of the protein (or its binding partner or ligand) in biological fluids; and as markers for tissues in which the corresponding protein is preferentially expressed (either constitutively or at a particular stage of tissue differentiation or development or in a disease state). Where the protein binds or potentially binds to another protein or ligand (such as, for example, in a enzyme-effector protein interaction or enzyme-ligand interaction), the protein can be used to identify the binding partner/ligand so as to develop a system to identify inhibitors of the binding interaction. Any or all of these uses are capable of being developed into reagent grade or kit format for commercialization as commercial products.

[0050] Methods for performing the uses listed above are well known to those skilled in the art. References disclosing such methods include “Molecular Cloning: A Laboratory Manual”, 2d ed., Cold Spring Harbor Laboratory Press, Sambrook, J., E. F. Fritsch and T. Maniatis eds., 1989, and “Methods in Enzymology: Guide to Molecular Cloning Techniques”, Academic Press, Berger, S. L. and A. R. Kimmel eds., 1987.

[0051] The potential uses of the peptides of the present invention are based primarily on the source of the protein as well as the class/action of the protein. For example, enzymes isolated from humans and their human/mammalian orthologs serve as targets for identifying agents for use in mammalian therapeutic applications, e.g. a human drug, particularly in modulating a biological or pathological response in a cell or tissue that expresses the enzyme. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus. A large percentage of pharmaceutical agents are being developed that modulate the activity of enzyme proteins, particularly members of the lanosterol synthase subfamily (see Background of the Invention). The structural and functional information provided in the Background and Figures provide specific and substantial uses for the molecules of the present invention, particularly in combination with the expression information provided in FIG. 1. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. Such uses can readily be determined using the information provided herein, that which is known in the art, and routine experimentation.

[0052] The proteins of the present invention (including variants and fragments that may have been disclosed prior to the present invention) are useful for biological assays related to enzymes that are related to members of the lanosterol synthase subfamily. Such assays involve any of the known enzyme functions or activities or properties useful for diagnosis and treatment of enzyme-related conditions that are specific for the subfamily of enzymes that the one of the present invention belongs to, particularly in cells and tissues that express the enzyme. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus.

[0053] The proteins of the present invention are also useful in drug screening assays, in cell-based or cell-free systems. Cell-based systems can be native, i.e., cells that normally express the enzyme, as a biopsy or expanded in cell culture. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. In an alternate embodiment, cell-based assays involve recombinant host cells expressing the enzyme protein.

[0054] The polypeptides can be used to identify compounds that modulate enzyme activity of the protein in its natural state or an altered form that causes a specific disease or pathology associated with the enzyme. Both the enzymes of the present invention and appropriate variants and fragments can be used in high-throughput screens to assay candidate compounds for the ability to bind to the enzyme. These compounds can be further screened against a functional enzyme to determine the effect of the compound on the enzyme activity. Further, these compounds can be tested in animal or invertebrate systems to determine activity/effectiveness. Compounds can be identified that activate (agonist) or inactivate (antagonist) the enzyme to a desired degree.

[0055] Further, the proteins of the present invention can be used to screen a compound for the ability to stimulate or inhibit interaction between the enzyme protein and a molecule that normally interacts with the enzyme protein, e.g. a substrate or a component of the signal pathway that the enzyme protein normally interacts (for example, another enzyme). Such assays typically include the steps of combining the enzyme protein with a candidate compound under conditions that allow the enzyme protein, or fragment, to interact with the target molecule, and to detect the formation of a complex between the protein and the target or to detect the biochemical consequence of the interaction with the enzyme protein and the target, such as any of the associated effects of signal transduction such as protein phosphorylation, cAMP turnover, and adenylate cyclase activation, etc.

[0056] Candidate compounds include, for example, 1) peptides such as soluble peptides, including Ig-tailed fusion peptides and members of random peptide libraries (see, e.g., Lam et al., Nature 354:82-84 (1991); Houghten et al., Nature 354:84-86 (1991)) and combinatorial chemistry-derived molecular libraries made of D- and/or L-configuration amino acids; 2) phosphopeptides (e.g., members of random and partially degenerate, directed phosphopeptide libraries, see, e.g., Songyang et al., Cell 72:767-778 (1993)); 3) antibodies (e.g., polyclonal, monoclonal, humanized, anti-idiotypic, chimeric, and single chain antibodies as well as Fab, F(ab)₂, Fab expression library fragments, and epitope-binding fragments of antibodies); and 4) small organic and inorganic molecules (e.g., molecules obtained from combinatorial and natural product libraries).

[0057] One candidate compound is a soluble fragment of the receptor that competes for substrate binding. Other candidate compounds include mutant enzymes or appropriate fragments containing mutations that affect enzyme function and thus compete for substrate. Accordingly, a fragment that competes for substrate, for example with a higher affinity, or a fragment that binds substrate but does not allow release, is encompassed by the invention.

[0058] The invention further includes other end point assays to identify compounds that modulate (stimulate or inhibit) enzyme activity. The assays typically involve an assay of events in the signal transduction pathway that indicate enzyme activity. Thus, the phosphorylation of a substrate, activation of a protein, a change in the expression of genes that are up- or down-regulated in response to the enzyme protein dependent signal cascade can be assayed.

[0059] Any of the biological or biochemical functions mediated by the enzyme can be used as an endpoint assay. These include all of the biochemical or biochemical/biological events described herein, in the references cited herein, incorporated by reference for these endpoint assay targets, and other functions known to those of ordinary skill in the art or that can be readily identified using the information provided in the Figures, particularly FIG. 2. Specifically, a biological function of a cell or tissues that expresses the enzyme can be assayed. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus.

[0060] Binding and/or activating compounds can also be screened by using chimeric enzyme proteins in which the amino terminal extracellular domain, or parts thereof, the entire transmembrane domain or subregions, such as any of the seven transmembrane segments or any of the intracellular or extracellular loops and the carboxy terminal intracellular domain, or parts thereof, can be replaced by heterologous domains or subregions. For example, a substrate-binding region can be used that interacts with a different substrate then that which is recognized by the native enzyme. Accordingly, a different set of signal transduction components is available as an end-point assay for activation. This allows for assays to be performed in other than the specific host cell from which the enzyme is derived.

[0061] The proteins of the present invention are also useful in competition binding assays in methods designed to discover compounds that interact with the enzyme (e.g. binding partners and/or ligands). Thus, a compound is exposed to a enzyme polypeptide under conditions that allow the compound to bind or to otherwise interact with the polypeptide. Soluble enzyme polypeptide is also added to the mixture. If the test compound interacts with the soluble enzyme polypeptide, it decreases the amount of complex formed or activity from the enzyme target. This type of assay is particularly useful in cases in which compounds are sought that interact with specific regions of the enzyme. Thus, the soluble polypeptide that competes with the target enzyme region is designed to contain peptide sequences corresponding to the region of interest.

[0062] To perform cell free drug screening assays, it is sometimes desirable to immobilize either the enzyme protein, or fragment, or its target molecule to facilitate separation of complexes from uncomplexed forms of one or both of the proteins, as well as to accommodate automation of the assay.

[0063] Techniques for immobilizing proteins on matrices can be used in the drug screening assays. In one embodiment, a fusion protein can be provided which adds a domain that allows the protein to be bound to a matrix. For example, glutathione-S-transferase fusion proteins can be adsorbed onto glutathione sepharose beads (Sigma Chemical, St. Louis, Mo.) or glutathione derivatized microtitre plates, which are then combined with the cell lysates (e.g., ³⁵S-labeled) and the candidate compound, and the mixture incubated under conditions conducive to complex formation (e.g., at physiological conditions for salt and pH). Following incubation, the beads are washed to remove any unbound label, and the matrix immobilized and radiolabel determined directly, or in the supernatant after the complexes are dissociated. Alternatively, the complexes can be dissociated from the matrix, separated by SDS-PAGE, and the level of enzyme-binding protein found in the bead fraction quantitated from the gel using standard electrophoretic techniques. For example, either the polypeptide or its target molecule can be immobilized utilizing conjugation of biotin and streptavidin using techniques well known in the art. Alternatively, antibodies reactive with the protein but which do not interfere with binding of the protein to its target molecule can be derivatized to the wells of the plate, and the protein trapped in the wells by antibody conjugation. Preparations of a enzyme-binding protein and a candidate compound are incubated in the enzyme protein-presenting wells and the amount of complex trapped in the well can be quantitated. Methods for detecting such complexes, in addition to those described above for the GST-immobilized complexes, include immunodetection of complexes using antibodies reactive with the enzyme protein target molecule, or which are reactive with enzyme protein and compete with the target molecule, as well as enzyme-linked assays which rely on detecting an enzymatic activity associated with the target molecule.

[0064] Agents that modulate one of the enzymes of the present invention can be identified using one or more of the above assays, alone or in combination. It is generally preferable to use a cell-based or cell free system first and then confirm activity in an animal or other model system. Such model systems are well known in the art and can readily be employed in this context.

[0065] Modulators of enzyme protein activity identified according to these drug screening assays can be used to treat a subject with a disorder mediated by the enzyme pathway, by treating cells or tissues that express the enzyme. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. These methods of treatment include the steps of administering a modulator of enzyme activity in a pharmaceutical composition to a subject in need of such treatment, the modulator being identified as described herein.

[0066] In yet another aspect of the invention, the enzyme proteins can be used as “bait proteins” in a two-hybrid assay or three-hybrid assay (see, e.g., U.S. Pat. No. 5,283,317; Zervos et al. (1993) Cell 72:223-232; Madura et al. (1993) J. Biol. Chem. 268:12046-12054; Bartel et al. (1993) Biotechniques 14:920-924; Iwabuchi et al. (1993) Oncogene 8:1693-1696; and Brent WO94/10300), to identify other proteins, which bind to or interact with the enzyme and are involved in enzyme activity. Such enzyme-binding proteins are also likely to be involved in the propagation of signals by the enzyme proteins or enzyme targets as, for example, downstream elements of a enzyme-mediated signaling pathway. Alternatively, such enzyme-binding proteins are likely to be enzyme inhibitors.

[0067] The two-hybrid system is based on the modular nature of most transcription factors, which consist of separable DNA-binding and activation domains. Briefly, the assay utilizes two different DNA constructs. In one construct, the gene that codes for a enzyme protein is fused to a gene encoding the DNA binding domain of a known transcription factor (e.g., GALA). In the other construct, a DNA sequence, from a library of DNA sequences, that encodes an unidentified protein (“prey” or “sample”) is fused to a gene that codes for the activation domain of the known transcription factor. If the “bait” and the “prey” proteins are able to interact, in vivo, forming a enzyme-dependent complex, the DNA-binding and activation domains of the transcription factor are brought into close proximity. This proximity allows transcription of a reporter gene (e.g., LacZ) which is operably linked to a transcriptional regulatory site responsive to the transcription factor. Expression of the reporter gene can be detected and cell colonies containing the functional transcription factor can be isolated and used to obtain the cloned gene which encodes the protein which interacts with the enzyme protein.

[0068] This invention further pertains to novel agents identified by the above-described screening assays. Accordingly, it is within the scope of this invention to further use an agent identified as described herein in an appropriate animal model. For example, an agent identified as described herein (e.g., a enzyme-modulating agent, an antisense enzyme nucleic acid molecule, a enzyme-specific antibody, or a enzyme-binding partner) can be used in an animal or other model to determine the efficacy, toxicity, or side effects of treatment with such an agent. Alternatively, an agent identified as described herein can be used in an animal or other model to determine the mechanism of action of such an agent. Furthermore, this invention pertains to uses of novel agents identified by the above-described screening assays for treatments as described herein.

[0069] The enzyme proteins of the present invention are also useful to provide a target for diagnosing a disease or predisposition to disease mediated by the peptide. Accordingly, the invention provides methods for detecting the presence, or levels of, the protein (or encoding mRNA) in a cell, tissue, or organism. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. The method involves contacting a biological sample with a compound capable of interacting with the enzyme protein such that the interaction can be detected. Such an assay can be provided in a single detection format or a multi-detection format such as an antibody chip array.

[0070] One agent for detecting a protein in a sample is an antibody capable of selectively binding to protein. A biological sample includes tissues, cells and biological fluids isolated from a subject, as well as tissues, cells and fluids present within a subject.

[0071] The peptides of the present invention also provide targets for diagnosing active protein activity, disease, or predisposition to disease, in a patient having a variant peptide, particularly activities and conditions that are known for other members of the family of proteins to which the present one belongs. Thus, the peptide can be isolated from a biological sample and assayed for the presence of a genetic mutation that results in aberrant peptide. This includes amino acid substitution, deletion, insertion, rearrangement, (as the result of aberrant splicing events), and inappropriate post-translational modification. Analytic methods include altered electrophoretic mobility, altered tryptic peptide digest, altered enzyme activity in cell-based or cell-free assay, alteration in substrate or antibody-binding pattern, altered isoelectric point, direct amino acid sequencing, and any other of the known assay techniques useful for detecting mutations in a protein. Such an assay can be provided in a single detection format or a multi-detection format such as an antibody chip array.

[0072] In vitro techniques for detection of peptide include enzyme linked immunosorbent assays (ELISAs), Western blots, immunoprecipitations and immunofluorescence using a detection reagent, such as an antibody or protein binding agent. Alternatively, the peptide can be detected in vivo in a subject by introducing into the subject a labeled anti-peptide antibody or other types of detection agent. For example, the antibody can be labeled with a radioactive marker whose presence and location in a subject can be detected by standard imaging techniques. Particularly useful are methods that detect the allelic variant of a peptide expressed in a subject and methods which detect fragments of a peptide in a sample.

[0073] The peptides are also useful in pharmacogenomic analysis. Pharmacogenomics deal with clinically significant hereditary variations in the response to drugs due to altered drug disposition and abnormal action in affected persons. See, e.g., Eichelbaum, M. (Clin. Exp. Pharmacol. Physiol. 23(10-11):983-985 (1996)), and Linder, M. W. (Clin. Chem. 43(2):254-266 (1997)). The clinical outcomes of these variations result in severe toxicity of therapeutic drugs in certain individuals or therapeutic failure of drugs in certain individuals as a result of individual variation in metabolism. Thus, the genotype of the individual can determine the way a therapeutic compound acts on the body or the way the body metabolizes the compound. Further, the activity of drug metabolizing enzymes effects both the intensity and duration of drug action. Thus, the pharmacogenomics of the individual permit the selection of effective compounds and effective dosages of such compounds for prophylactic or therapeutic treatment based on the individual's genotype. The discovery of genetic polymorphisms in some drug metabolizing enzymes has explained why some patients do not obtain the expected drug effects, show an exaggerated drug effect, or experience serious toxicity from standard drug dosages. Polymorphisms can be expressed in the phenotype of the extensive metabolizer and the phenotype of the poor metabolizer. Accordingly, genetic polymorphism may lead to allelic protein variants of the enzyme protein in which one or more of the enzyme functions in one population is different from those in another population. The peptides thus allow a target to ascertain a genetic predisposition that can affect treatment modality. Thus, in a ligand-based treatment, polymorphism may give rise to amino terminal extracellular domains and/or other substrate-binding regions that are more or less active in substrate binding, and enzyme activation. Accordingly, substrate dosage would necessarily be modified to maximize the therapeutic effect within a given population containing a polymorphism. As an alternative to genotyping, specific polymorphic peptides could be identified.

[0074] The peptides are also useful for treating a disorder characterized by an absence of, inappropriate, or unwanted expression of the protein. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. Accordingly, methods for treatment include the use of the enzyme protein or fragments.

[0075] Antibodies

[0076] The invention also provides antibodies that selectively bind to one of the peptides of the present invention, a protein comprising such a peptide, as well as variants and fragments thereof. As used herein, an antibody selectively binds a target peptide when it binds the target peptide and does not significantly bind to unrelated proteins. An antibody is still considered to selectively bind a peptide even if it also binds to other proteins that are not substantially homologous with the target peptide so long as such proteins share homology with a fragment or domain of the peptide target of the antibody. In this case, it would be understood that antibody binding to the peptide is still selective despite some degree of cross-reactivity.

[0077] As used herein, an antibody is defined in terms consistent with that recognized within the art: they are multi-subunit proteins produced by a mammalian organism in response to an antigen challenge. The antibodies of the present invention include polyclonal antibodies and monoclonal antibodies, as well as fragments of such antibodies, including, but not limited to, Fab or F(ab′)₂, and Fv fragments.

[0078] Many methods are known for generating and/or identifying antibodies to a given target peptide. Several such methods are described by Harlow, Antibodies, Cold Spring Harbor Press, (1989).

[0079] In general, to generate antibodies, an isolated peptide is used as an immunogen and is administered to a mammalian organism, such as a rat, rabbit or mouse. The full-length protein, an antigenic peptide fragment or a fusion protein can be used. Particularly important fragments are those covering functional domains, such as the domains identified in FIG. 2, and domain of sequence homology or divergence amongst the family, such as those that can readily be identified using protein alignment methods and as presented in the Figures.

[0080] Antibodies are preferably prepared from regions or discrete fragments of the enzyme proteins. Antibodies can be prepared from any region of the peptide as described herein. However, preferred regions will include those involved in function/activity and/or enzyme/binding partner interaction. FIG. 2 can be used to identify particularly important regions while sequence alignment can be used to identify conserved and unique sequence fragments.

[0081] An antigenic fragment will typically comprise at least 8 contiguous amino acid residues. The antigenic peptide can comprise, however, at least 10, 12, 14, 16 or more amino acid residues. Such fragments can be selected on a physical property, such as fragments correspond to regions that are located on the surface of the protein, e.g., hydrophilic regions or can be selected based on sequence uniqueness (see FIG. 2).

[0082] Detection on an antibody of the present invention can be facilitated by coupling (i.e., physically linking) the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, and radioactive materials. Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, β-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin, and examples of suitable radioactive material include ¹²⁵I, ¹³¹I, ³⁵S or ³H.

[0083] Antibody Uses

[0084] The antibodies can be used to isolate one of the proteins of the present invention by standard techniques, such as affinity chromatography or immunoprecipitation. The antibodies can facilitate the purification of the natural protein from cells and recombinantly produced protein expressed in host cells. In addition, such antibodies are useful to detect the presence of one of the proteins of the present invention in cells or tissues to determine the pattern of expression of the protein among various tissues in an organism and over the course of normal development. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus. Further, such antibodies can be used to detect protein in situ, in vitro, or in a cell lysate or supernatant in order to evaluate the abundance and pattern of expression. Also, such antibodies can be used to assess abnormal tissue distribution or abnormal expression during development or progression of a biological condition. Antibody detection of circulating fragments of the full length protein can be used to identify turnover.

[0085] Further, the antibodies can be used to assess expression in disease states such as in active stages of the disease or in an individual with a predisposition toward disease related to the protein's function. When a disorder is caused by an inappropriate tissue distribution, developmental expression, level of expression of the protein, or expressed/processed form, the antibody can be prepared against the normal protein. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. If a disorder is characterized by a specific mutation in the protein, antibodies specific for this mutant protein can be used to assay for the presence of the specific mutant protein.

[0086] The antibodies can also be used to assess normal and aberrant subcellular localization of cells in the various tissues in an organism. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. The diagnostic uses can be applied, not only in genetic testing, but also in monitoring a treatment modality. Accordingly, where treatment is ultimately aimed at correcting expression level or the presence of aberrant sequence and aberrant tissue distribution or developmental expression, antibodies directed against the protein or relevant fragments can be used to monitor therapeutic efficacy.

[0087] Additionally, antibodies are useful in pharmacogenomic analysis. Thus, antibodies prepared against polymorphic proteins can be used to identify individuals that require modified treatment modalities. The antibodies are also useful as diagnostic tools as an immunological marker for aberrant protein analyzed by electrophoretic mobility, isoelectric point, tryptic peptide digest, and other physical assays known to those in the art.

[0088] The antibodies are also useful for tissue typing. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. Thus, where a specific protein has been correlated with expression in a specific tissue, antibodies that are specific for this protein can be used to identify a tissue type.

[0089] The antibodies are also useful for inhibiting protein function, for example, blocking the binding of the enzyme peptide to a binding partner such as a substrate. These uses can also be applied in a therapeutic context in which treatment involves inhibiting the protein's function. An antibody can be used, for example, to block binding, thus modulating (agonizing or antagonizing) the peptides activity. Antibodies can be prepared against specific fragments containing sites required for function or against intact protein that is associated with a cell or cell membrane. See FIG. 2 for structural information relating to the proteins of the present invention.

[0090] The invention also encompasses kits for using antibodies to detect the presence of a protein in a biological sample. The kit can comprise antibodies such as a labeled or labelable antibody and a compound or agent for detecting protein in a biological sample; means for determining the amount of protein in the sample; means for comparing the amount of protein in the sample with a standard; and instructions for use. Such a kit can be supplied to detect a single protein or epitope or can be configured to detect one of a multitude of epitopes, such as in an antibody detection array. Arrays are described in detail below for nuleic acid arrays and similar methods have been developed for antibody arrays.

[0091] Nucleic Acid Molecules

[0092] The present invention further provides isolated nucleic acid molecules that encode a enzyme peptide or protein of the present invention (cDNA, transcript and genomic sequence). Such nucleic acid molecules will consist of, consist essentially of, or comprise a nucleotide sequence that encodes one of the enzyme peptides of the present invention, an allelic variant thereof, or an ortholog or paralog thereof.

[0093] As used herein, an “isolated” nucleic acid molecule is one that is separated from other nucleic acid present in the natural source of the nucleic acid. Preferably, an “isolated” nucleic acid is free of sequences which naturally flank the nucleic acid (i.e., sequences located at the 5′ and 3′ ends of the nucleic acid) in the genomic DNA of the organism from which the nucleic acid is derived. However, there can be some flanking nucleotide sequences, for example up to about 5 KB, 4 KB, 3 KB, 2 KB, or 1 KB or less, particularly contiguous peptide encoding sequences and peptide encoding sequences within the same gene but separated by introns in the genomic sequence. The important point is that the nucleic acid is isolated from remote and unimportant flanking sequences such that it can be subjected to the specific manipulations described herein such as recombinant expression, preparation of probes and primers, and other uses specific to the nucleic acid sequences.

[0094] Moreover, an “isolated” nucleic acid molecule, such as a transcript/cDNA molecule, can be substantially free of other cellular material, or culture medium when produced by recombinant techniques, or chemical precursors or other chemicals when chemically synthesized. However, the nucleic acid molecule can be fused to other coding or regulatory sequences and still be considered isolated.

[0095] For example, recombinant DNA molecules contained in a vector are considered isolated. Further examples of isolated DNA molecules include recombinant DNA molecules maintained in heterologous host cells or purified (partially or substantially) DNA molecules in solution. Isolated RNA molecules include in vivo or in vitro RNA transcripts of the isolated DNA molecules of the present invention. Isolated nucleic acid molecules according to the present invention further include such molecules produced synthetically.

[0096] Accordingly, the present invention provides nucleic acid molecules that consist of the nucleotide sequence shown in FIG. 1 or 3 (SEQ ID NO:1, transcript sequence and SEQ ID NO:3, genomic sequence), or any nucleic acid molecule that encodes the protein provided in FIG. 2, SEQ ID NO:2. A nucleic acid molecule consists of a nucleotide sequence when the nucleotide sequence is the complete nucleotide sequence of the nucleic acid molecule.

[0097] The present invention further provides nucleic acid molecules that consist essentially of the nucleotide sequence shown in FIG. 1 or 3 (SEQ ID NO:1, transcript sequence and SEQ ID NO:3, genomic sequence), or any nucleic acid molecule that encodes the protein provided in FIG. 2, SEQ ID NO:2. A nucleic acid molecule consists essentially of a nucleotide sequence when such a nucleotide sequence is present with only a few additional nucleic acid residues in the final nucleic acid molecule.

[0098] The present invention further provides nucleic acid molecules that comprise the nucleotide sequences shown in FIG. 1 or 3 (SEQ ID NO:1, transcript sequence and SEQ ID NO:3, genomic sequence), or any nucleic acid molecule that encodes the protein provided in FIG. 2, SEQ ID NO:2. A nucleic acid molecule comprises a nucleotide sequence when the nucleotide sequence is at least part of the final nucleotide sequence of the nucleic acid molecule. In such a fashion, the nucleic acid molecule can be only the nucleotide sequence or have additional nucleic acid residues, such as nucleic acid residues that are naturally associated with it or heterologous nucleotide sequences. Such a nucleic acid molecule can have a few additional nucleotides or can comprises several hundred or more additional nucleotides. A brief description of how various types of these nucleic acid molecules can be readily made/isolated is provided below.

[0099] In FIGS. 1 and 3, both coding and non-coding sequences are provided. Because of the source of the present invention, humans genomic sequence (FIG. 3) and cDNA/transcript sequences (FIG. 1), the nucleic acid molecules in the Figures will contain genomic intronic sequences, 5′ and 3′ non-coding sequences, gene regulatory regions and non-coding intergenic sequences. In general such sequence features are either noted in FIGS. 1 and 3 or can readily be identified using computational tools known in the art. As discussed below, some of the non-coding regions, particularly gene regulatory elements such as promoters, are useful for a variety of purposes, e.g. control of heterologous gene expression, target for identifying gene activity modulating compounds, and are particularly claimed as fragments of the genomic sequence provided herein.

[0100] The isolated nucleic acid molecules can encode the mature protein plus additional amino or carboxyl-terminal amino acids, or amino acids interior to the mature peptide (when the mature form has more than one peptide chain, for instance). Such sequences may play a role in processing of a protein from precursor to a mature form, facilitate protein trafficking, prolong or shorten protein half-life or facilitate manipulation of a protein for assay or production, among other things. As generally is the case in situ, the additional amino acids may be processed away from the mature protein by cellular enzymes.

[0101] As mentioned above, the isolated nucleic acid molecules include, but are not limited to, the sequence encoding the enzyme peptide alone, the sequence encoding the mature peptide and additional coding sequences, such as a leader or secretory sequence (e.g., a pre-pro or pro-protein sequence), the sequence encoding the mature peptide, with or without the additional coding sequences, plus additional non-coding sequences, for example introns and non-coding 5′ and 3′ sequences such as transcribed but non-translated sequences that play a role in transcription, mRNA processing (including splicing and polyadenylation signals), ribosome binding and stability of mRNA. In addition, the nucleic acid molecule may be fused to a marker sequence encoding, for example, a peptide that facilitates purification.

[0102] Isolated nucleic acid molecules can be in the form of RNA, such as mRNA, or in the form DNA, including cDNA and genormic DNA obtained by cloning or produced by chemical synthetic techniques or by a combination thereof. The nucleic acid, especially DNA, can be double-stranded or single-stranded. Single-stranded nucleic acid can be the coding strand (sense strand) or the non-coding strand (anti-sense strand).

[0103] The invention further provides nucleic acid molecules that encode fragments of the peptides of the present invention as well as nucleic acid molecules that encode obvious variants of the enzyme proteins of the present invention that are described above. Such nucleic acid molecules may be naturally occurring, such as allelic variants (same locus), paralogs (different locus), and orthologs (different organism), or may be constructed by recombinant DNA methods or by chemical synthesis. Such non-naturally occurring variants may be made by mutagenesis techniques, including those applied to nucleic acid molecules, cells, or organisms. Accordingly, as discussed above, the variants can contain nucleotide substitutions, deletions, inversions and insertions. Variation can occur in either or both the coding and non-coding regions. The variations can produce both conservative and non-conservative amino acid substitutions.

[0104] The present invention further provides non-coding fragments of the nucleic acid molecules provided in FIGS. 1 and 3. Preferred non-coding fragments include, but are not limited to, promoter sequences, enhancer sequences, gene modulating sequences and gene termination sequences. Such fragments are useful in controlling heterologous gene expression and in developing screens to identify gene-modulating agents. A promoter can readily be identified as being 5′ to the ATG start site in the genomic sequence provided in FIG. 3.

[0105] A fragment comprises a contiguous nucleotide sequence greater than 12 or more nucleotides. Further, a fragment could at least 30, 40, 50, 100, 250 or 500 nucleotides in length. The length of the fragment will be based on its intended use. For example, the fragment can encode epitope bearing regions of the peptide, or can be useful as DNA probes and primers. Such fragments can be isolated using the known nucleotide sequence to synthesize an oligonucleotide probe. A labeled probe can then be used to screen a cDNA library, genomic DNA library, or mRNA to isolate nucleic acid corresponding to the coding region. Further, primers can be used in PCR reactions to clone specific regions of gene.

[0106] A probe/primer typically comprises substantially a purified oligonucleotide or oligonucleotide pair. The oligonucleotide typically comprises a region of nucleotide sequence that hybridizes under stringent conditions to at least about 12, 20, 25, 40, 50 or more consecutive nucleotides.

[0107] Orthologs, homologs, and allelic variants can be identified using methods well known in the art. As described in the Peptide Section, these variants comprise a nucleotide sequence encoding a peptide that is typically 60-70%, 70-80%, 80-90%, and more typically at least about 90-95% or more homologous to the nucleotide sequence shown in the Figure sheets or a fragment of this sequence. Such nucleic acid molecules can readily be identified as being able to hybridize under moderate to stringent conditions, to the nucleotide sequence shown in the Figure sheets or a fragment of the sequence. Allelic variants can readily be determined by genetic locus of the encoding gene. The gene encoding the novel enzyme of the present invention is located on a genome component that has been mapped to human chromosome 21 (as indicated in FIG. 3), which is supported by multiple lines of evidence, such as STS and BAC map data.

[0108]FIG. 3 provides information on SNPs that have been found in the gene encoding the enzyme of the present invention. SNPs were identified at 56 different nucleotide positions, including a non-synonymous coding SNP at position 36001 (protein position 631). Changes in the amino acid sequence caused by these SNPs is indicated in FIG. 3 and can readily be determined using the universal genetic code and the protein sequence provided in FIG. 2 as a reference. Some of these SNPs, located outside the ORF and in introns, may affect gene transcription.

[0109] As used herein, the term “hybridizes under stringent conditions” is intended to describe conditions for hybridization and washing under which nucleotide sequences encoding a peptide at least 60-70% homologous to each other typically remain hybridized to each other. The conditions can be such that sequences at least about 60%, at least about 70%, or at least about 80% or more homologous to each other typically remain hybridized to each other. Such stringent conditions are known to those skilled in the art and can be found in Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1-6.3.6. One example of stringent hybridization conditions are hybridization in 6× sodium chloride/sodium citrate (SSC) at about 45C, followed by one or more washes in 0.2×SSC, 0.1% SDS at 50-65C. Examples of moderate to low stringency hybridization conditions are well known in the art.

[0110] Nucleic Acid Molecule Uses

[0111] The nucleic acid molecules of the present invention are useful for probes, primers, chemical intermediates, and in biological assays. The nucleic acid molecules are useful as a hybridization probe for messenger RNA, transcript/cDNA and genomic DNA to isolate full-length cDNA and genomic clones encoding the peptide described in FIG. 2 and to isolate cDNA and genomic clones that correspond to variants (alleles, orthologs, etc.) producing the same or related peptides shown in FIG. 2. As illustrated in FIG. 3, SNPs were identified at 56 different nucleotide positions.

[0112] The probe can correspond to any sequence along the entire length of the nucleic acid molecules provided in the Figures. Accordingly, it could be derived from 5′ noncoding regions, the coding region, and 3′ noncoding regions. However, as discussed, fragments are not to be construed as encompassing fragments disclosed prior to the present invention.

[0113] The nucleic acid molecules are also useful as primers for PCR to amplify any given region of a nucleic acid molecule and are useful to synthesize antisense molecules of desired length and sequence.

[0114] The nucleic acid molecules are also useful for constructing recombinant vectors. Such vectors include expression vectors that express a portion of, or all of, the peptide sequences. Vectors also include insertion vectors, used to integrate into another nucleic acid molecule sequence, such as into the cellular genome, to alter in situ expression of a gene and/or gene product. For example, an endogenous coding sequence can be replaced via homologous recombination with all or part of the coding region containing one or more specifically introduced mutations.

[0115] The nucleic acid molecules are also useful for expressing antigenic portions of the proteins.

[0116] The nucleic acid molecules are also useful as probes for determining the chromosomal positions of the nucleic acid molecules by means of in situ hybridization methods. The gene encoding the novel enzyme of the present invention is located on a genome component that has been mapped to human chromosome 21 (as indicated in FIG. 3), which is supported by multiple lines of evidence, such as STS and BAC map data.

[0117] The nucleic acid molecules are also useful in making vectors containing the gene regulatory regions of the nucleic acid molecules of the present invention.

[0118] The nucleic acid molecules are also useful for designing ribozymes corresponding to all, or a part, of the mRNA produced from the nucleic acid molecules described herein.

[0119] The nucleic acid molecules are also useful for making vectors that express part, or all, of the peptides.

[0120] The nucleic acid molecules are also useful for constructing host cells expressing a part, or all, of the nucleic acid molecules and peptides.

[0121] The nucleic acid molecules are also useful for constructing transgenic animals expressing all, or a part, of the nucleic acid molecules and peptides.

[0122] The nucleic acid molecules are also useful as hybridization probes for determining the presence, level, form and distribution of nucleic acid expression. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus. Accordingly, the probes can be used to detect the presence of, or to determine levels of, a specific nucleic acid molecule in cells, tissues, and in organisms. The nucleic acid whose level is determined can be DNA or RNA. Accordingly, probes corresponding to the peptides described herein can be used to assess expression and/or gene copy number in a given cell, tissue, or organism. These uses are relevant for diagnosis of disorders involving an increase or decrease in enzyme protein expression relative to normal results.

[0123] In vitro techniques for detection of mRNA include Northern hybridizations and in situ hybridizations. In vitro techniques for detecting DNA includes Southern hybridizations and in situ hybridization.

[0124] Probes can be used as a part of a diagnostic test kit for identifying cells or tissues that express a enzyme protein, such as by measuring a level of a enzyme-encoding nucleic acid in a sample of cells from a subject e.g., mRNA or genomic DNA, or determining if a enzyme gene has been mutated. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus.

[0125] Nucleic acid expression assays are useful for drug screening to identify compounds that modulate enzyme nucleic acid expression.

[0126] The invention thus provides a method for identifying a compound that can be used to treat a disorder associated with nucleic acid expression of the enzyme gene, particularly biological and pathological processes that are mediated by the enzyme in cells and tissues that express it. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus. The method typically includes assaying the ability of the compound to modulate the expression of the enzyme nucleic acid and thus identifying a compound that can be used to treat a disorder characterized by undesired enzyme nucleic acid expression. The assays can be performed in cell-based and cell-free systems. Cell-based assays include cells naturally expressing the enzyme nucleic acid or recombinant cells genetically engineered to express specific nucleic acid sequences.

[0127] The assay for enzyme nucleic acid expression can involve direct assay of nucleic acid levels, such as mRNA levels, or on collateral compounds involved in the signal pathway. Further, the expression of genes that are up- or down-regulated in response to the enzyme protein signal pathway can also be assayed. In this embodiment the regulatory regions of these genes can be operably linked to a reporter gene such as luciferase.

[0128] Thus, modulators of enzyme gene expression can be identified in a method wherein a cell is contacted with a candidate compound and the expression of mRNA determined. The level of expression of enzyme mRNA in the presence of the candidate compound is compared to the level of expression of enzyme mRNA in the absence of the candidate compound. The candidate compound can then be identified as a modulator of nucleic acid expression based on this comparison and be used, for example to treat a disorder characterized by aberrant nucleic acid expression. When expression of mRNA is statistically significantly greater in the presence of the candidate compound than in its absence, the candidate compound is identified as a stimulator of nucleic acid expression. When nucleic acid expression is statistically significantly less in the presence of the candidate compound than in its absence, the candidate compound is identified as an inhibitor of nucleic acid expression.

[0129] The invention further provides methods of treatment, with the nucleic acid as a target, using a compound identified through drug screening as a gene modulator to modulate enzyme nucleic acid expression in cells and tissues that express the enzyme. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus. Modulation includes both up-regulation (i.e. activation or agonization) or down-regulation (suppression or antagonization) or nucleic acid expression.

[0130] Alternatively, a modulator for enzyme nucleic acid expression can be a small molecule or drug identified using the screening assays described herein as long as the drug or small molecule inhibits the enzyme nucleic acid expression in the cells and tissues that express the protein. Experimental data as provided in FIG. 1 indicates expression in humans in teratocarcinoma, ovary, uterus, muscle, brain, colon, and hippocampus.

[0131] The nucleic acid molecules are also useful for monitoring the effectiveness of modulating compounds on the expression or activity of the enzyme gene in clinical trials or in a treatment regimen. Thus, the gene expression pattern can serve as a barometer for the continuing effectiveness of treatment with the compound, particularly with compounds to which a patient can develop resistance. The gene expression pattern can also serve as a marker indicative of a physiological response of the affected cells to the compound. Accordingly, such monitoring would allow either increased administration of the compound or the administration of alternative compounds to which the patient has not become resistant. Similarly, if the level of nucleic acid expression falls below a desirable level, administration of the compound could be commensurately decreased.

[0132] The nucleic acid molecules are also useful in diagnostic assays for qualitative changes in enzyme nucleic acid expression, and particularly in qualitative changes that lead to pathology. The nucleic acid molecules can be used to detect mutations in enzyme genes and gene expression products such as mRNA. The nucleic acid molecules can be used as hybridization probes to detect naturally occurring genetic mutations in the enzyme gene and thereby to determine whether a subject with the mutation is at risk for a disorder caused by the mutation. Mutations include deletion, addition, or substitution of one or more nucleotides in the gene, chromosomal rearrangement, such as inversion or transposition, modification of genomic DNA, such as aberrant methylation patterns or changes in gene copy number, such as amplification. Detection of a mutated form of the enzyme gene associated with a dysfunction provides a diagnostic tool for an active disease or susceptibility to disease when the disease results from overexpression, underexpression, or altered expression of a enzyme protein.

[0133] Individuals carrying mutations in the enzyme gene can be detected at the nucleic acid level by a variety of techniques. FIG. 3 provides information on SNPs that have been found in the gene encoding the enzyme of the present invention. SNPs were identified at 56 different nucleotide positions, including a non-synonymous coding SNP at position 36001 (protein position 631). Changes in the amino acid sequence caused by these SNPs is indicated in FIG. 3 and can readily be determined using the universal genetic code and the protein sequence provided in FIG. 2 as a reference. Some of these SNPs, located outside the ORF and in introns, may affect gene transcription. The gene encoding the novel enzyme of the present invention is located on a genome component that has been mapped to human chromosome 21 (as indicated in FIG. 3), which is supported by multiple lines of evidence, such as STS and BAC map data. Genomic DNA can be analyzed directly or can be amplified by using PCR prior to analysis. RNA or cDNA can be used in the same way. In some uses, detection of the mutation involves the use of a probe/primer in a polymerase chain reaction (PCR) (see, e.g. U.S. Pat. Nos. 4,683,195 and 4,683,202), such as anchor PCR or RACE PCR, or, alternatively, in a ligation chain reaction (LCR) (see, e.g., Landegran et al., Science 241:1077-1080 (1988); and Nakazawa et al., PNAS 91:360-364 (1994)), the latter of which can be particularly useful for detecting point mutations in the gene (see Abravaya et al., Nucleic Acids Res. 23:675-682 (1995)). This method can include the steps of collecting a sample of cells from a patient, isolating nucleic acid (e.g., genomic, mRNA or both) from the cells of the sample, contacting the nucleic acid sample with one or more primers which specifically hybridize to a gene under conditions such that hybridization and amplification of the gene (if present) occurs, and detecting the presence or absence of an amplification product, or detecting the size of the amplification product and comparing the length to a control sample. Deletions and insertions can be detected by a change in size of the amplified product compared to the normal genotype. Point mutations can be identified by hybridizing amplified DNA to normal RNA or antisense DNA sequences.

[0134] Alternatively, mutations in a enzyme gene can be directly identified, for example, by alterations in restriction enzyme digestion patterns determined by gel electrophoresis.

[0135] Further, sequence-specific ribozymes (U.S. Pat. No. 5,498,531) can be used to score for the presence of specific mutations by development or loss of a ribozyme cleavage site. Perfectly matched sequences can be distinguished from mismatched sequences by nuclease cleavage digestion assays or by differences in melting temperature.

[0136] Sequence changes at specific locations can also be assessed by nuclease protection assays such as RNase and S1 protection or the chemical cleavage method. Furthermore, sequence differences between a mutant enzyme gene and a wild-type gene can be determined by direct DNA sequencing. A variety of automated sequencing procedures can be utilized when performing the diagnostic assays (Naeve, C. W., (1995) Biotechniques 19:448), including sequencing by mass spectrometry (see, e.g., PCT International Publication No. WO 94/16101; Cohen et al., Adv. Chromatogr. 36:127-162 (1996); and Griffin et al., Appl. Biochem. Biotechnol. 38:147-159 (1993)).

[0137] Other methods for detecting mutations in the gene include methods in which protection from cleavage agents is used to detect mismatched bases in RNA/RNA or RNA/DNA duplexes (Myers et al., Science 230:1242 (1985)); Cotton et al., PNAS 85:4397 (1988); Saleeba et al., Meth. Enzymol. 217:286-295 (1992)), electrophoretic mobility of mutant and wild type nucleic acid is compared (Orita et al., PNAS 86:2766 (1989); Cotton et al., Mutat. Res. 285:125-144 (1993); and Hayashi et al., Genet. Anal. Tech. Appl. 9:73-79 (1992)), and movement of mutant or wild-type fragments in polyacrylamide gels containing a gradient of denaturant is assayed using denaturing gradient gel electrophoresis (Myers et al., Nature 313:495 (1985)). Examples of other techniques for detecting point mutations include selective oligonucleotide hybridization, selective amplification, and selective primer extension.

[0138] The nucleic acid molecules are also useful for testing an individual for a genotype that while not necessarily causing the disease nevertheless affects the treatment modality. Thus, the nucleic acid molecules can be used to study the relationship between an individual's genotype and the individual's response to a compound used for treatment (pharmacogenomic relationship). Accordingly, the nucleic acid molecules described herein can be used to assess the mutation content of the enzyme gene in an individual in order to select an appropriate compound or dosage regimen for treatment. FIG. 3 provides information on SNPs that have been found in the gene encoding the enzyme of the present invention. SNPs were identified at 56 different nucleotide positions, including a non-synonymous coding SNP at position 36001 (protein position 631). Changes in the amino acid sequence caused by these SNPs is indicated in FIG. 3 and can readily be determined using the universal genetic code and the protein sequence provided in FIG. 2 as a reference. Some of these SNPs, located outside the ORF and in introns, may affect gene transcription.

[0139] Thus nucleic acid molecules displaying genetic variations that affect treatment provide a diagnostic target that can be used to tailor treatment in an individual. Accordingly, the production of recombinant cells and animals containing these polymorphisms allow effective clinical design of treatment compounds and dosage regimens.

[0140] The nucleic acid molecules are thus useful as antisense constructs to control enzyme gene expression in cells, tissues, and organisms. A DNA antisense nucleic acid molecule is designed to be complementary to a region of the gene involved in transcription, preventing transcription and hence production of enzyme protein. An antisense RNA or DNA nucleic acid molecule would hybridize to the mRNA and thus block translation of mRNA into enzyme protein.

[0141] Alternatively, a class of antisense molecules can be used to inactivate mRNA in order to decrease expression of enzyme nucleic acid. Accordingly, these molecules can treat a disorder characterized by abnormal or undesired enzyme nucleic acid expression. This technique involves cleavage by means of ribozymes containing nucleotide sequences complementary to one or more regions in the mRNA that attenuate the ability of the mRNA to be translated. Possible regions include coding regions and particularly coding regions corresponding to the catalytic and other functional activities of the enzyme protein, such as substrate binding.

[0142] The nucleic acid molecules also provide vectors for gene therapy in patients containing cells that are aberrant in enzyme gene expression. Thus, recombinant cells, which include the patient's cells that have been engineered ex vivo and returned to the patient, are introduced into an individual where the cells produce the desired enzyme protein to treat the individual.

[0143] The invention also encompasses kits for detecting the presence of a enzyme nucleic acid in a biological sample. Experimental data as provided in FIG. 1 indicates that the enzymes of the present invention are expressed in humans in teratocarcinoma, ovary, uterus, muscle, brain, and colon, as indicated by virtual northern blot analysis. In addition, PCR-based tissue screening panels indicate expression in the hioppocampus. For example, the kit can comprise reagents such as a labeled or labelable nucleic acid or agent capable of detecting enzyme nucleic acid in a biological sample; means for determining the amount of enzyme nucleic acid in the sample; and means for comparing the amount of enzyme nucleic acid in the sample with a standard. The compound or agent can be packaged in a suitable container. The kit can further comprise instructions for using the kit to detect enzyme protein mRNA or DNA.

[0144] Nucleic Acid Arrays

[0145] The present invention further provides nucleic acid detection kits, such as arrays or microarrays of nucleic acid molecules that are based on the sequence information provided in FIGS. 1 and 3 (SEQ ID NOS: 1 and 3).

[0146] As used herein “Arrays” or “Microarrays” refers to an array of distinct polynucleotides or oligonucleotides synthesized on a substrate, such as paper, nylon or other type of membrane, filter, chip, glass slide, or any other suitable solid support. In one embodiment, the microarray is prepared and used according to the methods described in U.S. Pat. No. 5,837,832, Chee et al., PCT application WO95/11995 (Chee et al.), Lockhart, D. J. et al. (1996; Nat. Biotech. 14: 1675-1680) and Schena, M. et al. (1996; Proc. Natl. Acad. Sci. 93: 10614-10619), all of which are incorporated herein in their entirety by reference. In other embodiments, such arrays are produced by the methods described by Brown et al., U.S. Pat. No. 5,807,522.

[0147] The microarray or detection kit is preferably composed of a large number of unique, single-stranded nucleic acid sequences, usually either synthetic antisense oligonucleotides or fragments of cDNAs, fixed to a solid support. The oligonucleotides are preferably about 6-60 nucleotides in length, more preferably 15-30 nucleotides in length, and most preferably about 20-25 nucleotides in length. For a certain type of microarray or detection kit, it may be preferable to use oligonucleotides that are only 7-20 nucleotides in length. The microarray or detection kit may contain oligonucleotides that cover the known 5′, or 3′, sequence, sequential oligonucleotides which cover the full length sequence; or unique oligonucleotides selected from particular areas along the length of the sequence. Polynucleotides used in the microarray or detection kit may be oligonucleotides that are specific to a gene or genes of interest.

[0148] In order to produce oligonucleotides to a known sequence for a microarray or detection kit, the gene(s) of interest (or an ORF identified from the contigs of the present invention) is typically examined using a computer algorithm which starts at the 5′ or at the 3′ end of the nucleotide sequence. Typical algorithms will then identify oligomers of defined length that are unique to the gene, have a GC content within a range suitable for hybridization, and lack predicted secondary structure that may interfere with hybridization. In certain situations it may be appropriate to use pairs of oligonucleotides on a microarray or detection kit. The “pairs” will be identical, except for one nucleotide that preferably is located in the center of the sequence. The second oligonucleotide in the pair (mismatched by one) serves as a control. The number of oligonucleotide pairs may range from two to one million. The oligomers are synthesized at designated areas on a substrate using a light-directed chemical process. The substrate may be paper, nylon or other type of membrane, filter, chip, glass slide or any other suitable solid support.

[0149] In another aspect, an oligonucleotide may be synthesized on the surface of the substrate by using a chemical coupling procedure and an ink jet application apparatus, as described in PCT application WO95/251116 (Baldeschweiler et al.) which is incorporated herein in its entirety by reference. In another aspect, a “gridded” array analogous to a dot (or slot) blot may be used to arrange and link cDNA fragments or oligonucleotides to the surface of a substrate using a vacuum system, thermal, UV, mechanical or chemical bonding procedures. An array, such as those described above, may be produced by hand or by using available devices (slot blot or dot blot apparatus), materials (any suitable solid support), and machines (including robotic instruments), and may contain 8, 24, 96, 384, 1536, 6144 or more oligonucleotides, or any other number between two and one million which lends itself to the efficient use of commercially available instrumentation.

[0150] In order to conduct sample analysis using a microarray or detection kit, the RNA or DNA from a biological sample is made into hybridization probes. The mRNA is isolated, and cDNA is produced and used as a template to make antisense RNA (aRNA). The aRNA is amplified in the presence of fluorescent nucleotides, and labeled probes are incubated with the microarray or detection kit so that the probe sequences hybridize to complementary oligonucleotides of the microarray or detection kit. Incubation conditions are adjusted so that hybridization occurs with precise complementary matches or with various degrees of less complementarity. After removal of nonhybridized probes, a scanner is used to determine the levels and patterns of fluorescence. The scanned images are examined to determine degree of complementarity and the relative abundance of each oligonucleotide sequence on the microarray or detection kit. The biological samples may be obtained from any bodily fluids (such as blood, urine, saliva, phlegm, gastric juices, etc.), cultured cells, biopsies, or other tissue preparations. A detection system may be used to measure the absence, presence, and amount of hybridization for all of the distinct sequences simultaneously. This data may be used for large-scale correlation studies on the sequences, expression patterns, mutations, variants, or polymorphisms among samples.

[0151] Using such arrays, the present invention provides methods to identify the expression of the enzyme proteins/peptides of the present invention. In detail, such methods comprise incubating a test sample with one or more nucleic acid molecules and assaying for binding of the nucleic acid molecule with components within the test sample. Such assays will typically involve arrays comprising many genes, at least one of which is a gene of the present invention and or alleles of the enzyme gene of the present invention. FIG. 3 provides information on SNPs that have been found in the gene encoding the enzyme of the present invention. SNPs were identified at 56 different nucleotide positions, including a non-synonymous coding SNP at position 36001 (protein position 631). Changes in the amino acid sequence caused by these SNPs is indicated in FIG. 3 and can readily be determined using the universal genetic code and the protein sequence provided in FIG. 2 as a reference. Some of these SNPs, located outside the ORF and in introns, may affect gene transcription.

[0152] Conditions for incubating a nucleic acid molecule with a test sample vary. Incubation conditions depend on the format employed in the assay, the detection methods employed, and the type and nature of the nucleic acid molecule used in the assay. One skilled in the art will recognize that any one of the commonly available hybridization, amplification or array assay formats can readily be adapted to employ the novel fragments of the Human genome disclosed herein. Examples of such assays can be found in Chard, T, An Introduction to Radioimmunoassay and Related Techniques, Elsevier Science Publishers, Amsterdam, The Netherlands (1986); Bullock, G. R. et al., Techniques in Immunocytochemistry, Academic Press, Orlando, Fla. Vol. 1 (1982), Vol. 2 (1983), Vol. 3 (1985); Tijssen, P., Practice and Theory of Enzyme Immunoassays: Laboratory Techniques in Biochemistry and Molecular Biology, Elsevier Science Publishers, Amsterdam, The Netherlands (1985).

[0153] The test samples of the present invention include cells, protein or membrane extracts of cells. The test sample used in the above-described method will vary based on the assay format, nature of the detection method and the tissues, cells or extracts used as the sample to be assayed. Methods for preparing nucleic acid extracts or of cells are well known in the art and can be readily be adapted in order to obtain a sample that is compatible with the system utilized.

[0154] In another embodiment of the present invention, kits are provided which contain the necessary reagents to carry out the assays of the present invention.

[0155] Specifically, the invention provides a compartmentalized kit to receive, in close confinement, one or more containers which comprises: (a) a first container comprising one of the nucleic acid molecules that can bind to a fragment of the Human genome disclosed herein; and (b) one or more other containers comprising one or more of the following: wash reagents, reagents capable of detecting presence of a bound nucleic acid.

[0156] In detail, a compartmentalized kit includes any kit in which reagents are contained in separate containers. Such containers include small glass containers, plastic containers, strips of plastic, glass or paper, or arraying material such as silica. Such containers allows one to efficiently transfer reagents from one compartment to another compartment such that the samples and reagents are not cross-contaminated, and the agents or solutions of each container can be added in a quantitative fashion from one compartment to another. Such containers will include a container which will accept the test sample, a container which contains the nucleic acid probe, containers which contain wash reagents (such as phosphate buffered saline, Tris-buffers, etc.), and containers which contain the reagents used to detect the bound probe. One skilled in the art will readily recognize that the previously unidentified enzyme gene of the present invention can be routinely identified using the sequence information disclosed herein can be readily incorporated into one of the established kit formats which are well known in the art, particularly expression arrays.

[0157] Vectors/Host Cells

[0158] The invention also provides vectors containing the nucleic acid molecules described herein. The term “vector” refers to a vehicle, preferably a nucleic acid molecule, which can transport the nucleic acid molecules. When the vector is a nucleic acid molecule, the nucleic acid molecules are covalently linked to the vector nucleic acid. With this aspect of the invention, the vector includes a plasmid, single or double stranded phage, a single or double stranded RNA or DNA viral vector, or artificial chromosome, such as a BAC, PAC, YAC, OR MAC.

[0159] A vector can be maintained in the host cell as an extrachromosomal element where it replicates and produces additional copies of the nucleic acid molecules. Alternatively, the vector may integrate into the host cell genome and produce additional copies of the nucleic acid molecules when the host cell replicates.

[0160] The invention provides vectors for the maintenance (cloning vectors) or vectors for expression (expression vectors) of the nucleic acid molecules. The vectors can function in prokaryotic or eukaryotic cells or in both (shuttle vectors).

[0161] Expression vectors contain cis-acting regulatory regions that are operably linked in the vector to the nucleic acid molecules such that transcription of the nucleic acid molecules is allowed in a host cell. The nucleic acid molecules can be introduced into the host cell with a separate nucleic acid molecule capable of affecting transcription. Thus, the second nucleic acid molecule may provide a trans-acting factor interacting with the cis-regulatory control region to allow transcription of the nucleic acid molecules from the vector. Alternatively, a trans-acting factor may be supplied by the host cell. Finally, a trans-acting factor can be produced from the vector itself. It is understood, however, that in some embodiments, transcription and/or translation of the nucleic acid molecules can occur in a cell-free system.

[0162] The regulatory sequence to which the nucleic acid molecules described herein can be operably linked include promoters for directing mRNA transcription. These include, but are not limited to, the left promoter from bacteriophage λ, the lac, TRP, and TAC promoters from E. coli, the early and late promoters from SV40, the CMV immediate early promoter, the adenovirus early and late promoters, and retrovirus long-terminal repeats.

[0163] In addition to control regions that promote transcription, expression vectors may also include regions that modulate transcription, such as repressor binding sites and enhancers. Examples include the SV40 enhancer, the cytomegalovirus immediate early enhancer, polyoma enhancer, adenovirus enhancers, and retrovirus LTR enhancers.

[0164] In addition to containing sites for transcription initiation and control, expression vectors can also contain sequences necessary for transcription termination and, in the transcribed region a ribosome binding site for translation. Other regulatory control elements for expression include initiation and termination codons as well as polyadenylation signals. The person of ordinary skill in the art would be aware of the numerous regulatory sequences that are useful in expression vectors. Such regulatory sequences are described, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual. 2nd. ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., (1989).

[0165] A variety of expression vectors can be used to express a nucleic acid molecule. Such vectors include chromosomal, episomal, and virus-derived vectors, for example vectors derived from bacterial plasmids, from bacteriophage, from yeast episomes, from yeast chromosomal elements, including yeast artificial chromosomes, from viruses such as baculoviruses, papovaviruses such as SV40, Vaccinia viruses, adenoviruses, poxviruses, pseudorabies viruses, and retroviruses. Vectors may also be derived from combinations of these sources such as those derived from plasmid and bacteriophage genetic elements, e.g. cosmids and phagemids. Appropriate cloning and expression vectors for prokaryotic and eukaryotic hosts are described in Sambrook et al., Molecular Cloning: A Laboratory Manual. 2nd. ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., (1989).

[0166] The regulatory sequence may provide constitutive expression in one or more host cells (i.e. tissue specific) or may provide for inducible expression in one or more cell types such as by temperature, nutrient additive, or exogenous factor such as a hormone or other ligand. A variety of vectors providing for constitutive and inducible expression in prokaryotic and eukaryotic hosts are well known to those of ordinary skill in the art.

[0167] The nucleic acid molecules can be inserted into the vector nucleic acid by well-known methodology. Generally, the DNA sequence that will ultimately be expressed is joined to an expression vector by cleaving the DNA sequence and the expression vector with one or more restriction enzymes and then ligating the fragments together. Procedures for restriction enzyme digestion and ligation are well known to those of ordinary skill in the art.

[0168] The vector containing the appropriate nucleic acid molecule can be introduced into an appropriate host cell for propagation or expression using well-known techniques. Bacterial cells include, but are not limited to, E. coli, Streptomyces, and Salmonella typhimurium. Eukaryotic cells include, but are not limited to, yeast, insect cells such as Drosophila, animal cells such as COS and CHO cells, and plant cells.

[0169] As described herein, it may be desirable to express the peptide as a fusion protein. Accordingly, the invention provides fusion vectors that allow for the production of the peptides. Fusion vectors can increase the expression of a recombinant protein, increase the solubility of the recombinant protein, and aid in the purification of the protein by acting for example as a ligand for affinity purification. A proteolytic cleavage site may be introduced at the junction of the fusion moiety so that the desired peptide can ultimately be separated from the fusion moiety. Proteolytic enzymes include, but are not limited to, factor Xa, thrombin, and enteroenzyme. Typical fusion expression vectors include pGEX (Smith et al., Gene 67:31-40 (1988)), pMAL (New England Biolabs, Beverly, Mass.) and pRIT5 (Pharmacia, Piscataway, N.J.) which fuse glutathione S-transferase (GST), maltose E binding protein, or protein A, respectively, to the target recombinant protein. Examples of suitable inducible non-fusion E. coli expression vectors include pTrc (Amann et al., Gene 69:301-315 (1988)) and pET 11d (Studier et al., Gene Expression Technology: Methods in Enzymology 185:60-89 (1990)).

[0170] Recombinant protein expression can be maximized in host bacteria by providing a genetic background wherein the host cell has an impaired capacity to proteolytically cleave the recombinant protein. (Gottesman, S., Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990) 119-128). Alternatively, the sequence of the nucleic acid molecule of interest can be altered to provide preferential codon usage for a specific host cell, for example E. coli. (Wada et al., Nucleic Acids Res. 20:2111-2118 (1992)).

[0171] The nucleic acid molecules can also be expressed by expression vectors that are operative in yeast. Examples of vectors for expression in yeast e.g., S. cerevisiae include pYepSec1 (Baldari, et al., EMBO J. 6:229-234 (1987)), pMFa (Kurjan et al., Cell 30:933-943(1982)), pJRY88 (Schultz et al., Gene 54:113-123 (1987)), and pYES2 (Invitrogen Corporation, San Diego, Calif.).

[0172] The nucleic acid molecules can also be expressed in insect cells using, for example, baculovirus expression vectors. Baculovirus vectors available for expression of proteins in cultured insect cells (e.g., Sf 9 cells) include the pAc series (Smith et al., Mol. Cell Biol. 3:2156-2165 (1983)) and the pVL series (Lucklow et al., Virology 170:31-39 (1989)).

[0173] In certain embodiments of the invention, the nucleic acid molecules described herein are expressed in mammalian cells using mammalian expression vectors. Examples of mammalian expression vectors include pCDM8 (Seed, B. Nature 329:840(1987)) and pMT2PC (Kaufman et al., EMBO J. 6:187-195 (1987)).

[0174] The expression vectors listed herein are provided by way of example only of the well-known vectors available to those of ordinary skill in the art that would be useful to express the nucleic acid molecules. The person of ordinary skill in the art would be aware of other vectors suitable for maintenance propagation or expression of the nucleic acid molecules described herein. These are found for example in Sambrook, J., Fritsh, E. F., and Maniatis, T. Molecular Cloning: A Laboratory Manual. 2nd, ed., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989.

[0175] The invention also encompasses vectors in which the nucleic acid sequences described herein are cloned into the vector in reverse orientation, but operably linked to a regulatory sequence that permits transcription of antisense RNA. Thus, an antisense transcript can be produced to all, or to a portion, of the nucleic acid molecule sequences described herein, including both coding and non-coding regions. Expression of this antisense RNA is subject to each of the parameters described above in relation to expression of the sense RNA (regulatory sequences, constitutive or inducible expression, tissue-specific expression).

[0176] The invention also relates to recombinant host cells containing the vectors described herein. Host cells therefore include prokaryotic cells, lower eukaryotic cells such as yeast, other eukaryotic cells such as insect cells, and higher eukaryotic cells such as mammalian cells.

[0177] The recombinant host cells are prepared by introducing the vector constructs described herein into the cells by techniques readily available to the person of ordinary skill in the art. These include, but are not limited to, calcium phosphate transfection, DEAE-dextran-mediated transfection, cationic lipid-mediated transfection, electroporation, transduction, infection, lipofection, and other techniques such as those found in Sambrook, et al. (Molecular Cloning: A Laboratory Manual. 2nd, ed., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989).

[0178] Host cells can contain more than one vector. Thus, different nucleotide sequences can be introduced on different vectors of the same cell. Similarly, the nucleic acid molecules can be introduced either alone or with other nucleic acid molecules that are not related to the nucleic acid molecules such as those providing trans-acting factors for expression vectors. When more than one vector is introduced into a cell, the vectors can be introduced independently, co-introduced or joined to the nucleic acid molecule vector.

[0179] In the case of bacteriophage and viral vectors, these can be introduced into cells as packaged or encapsulated virus by standard procedures for infection and transduction. Viral vectors can be replication-competent or replication-defective. In the case in which viral replication is defective, replication will occur in host cells providing functions that complement the defects.

[0180] Vectors generally include selectable markers that enable the selection of the subpopulation of cells that contain the recombinant vector constructs. The marker can be contained in the same vector that contains the nucleic acid molecules described herein or may be on a separate vector. Markers include tetracycline or ampicillin-resistance genes for prokaryotic host cells and dihydrofolate reductase or neomycin resistance for eukaryotic host cells. However, any marker that provides selection for a phenotypic trait will be effective.

[0181] While the mature proteins can be produced in bacteria, yeast, mammalian cells, and other cells under the control of the appropriate regulatory sequences, cell-free transcription and translation systems can also be used to produce these proteins using RNA derived from the DNA constructs described herein.

[0182] Where secretion of the peptide is desired, which is difficult to achieve with multi-transmembrane domain containing proteins such as enzymes, appropriate secretion signals are incorporated into the vector. The signal sequence can be endogenous to the peptides or heterologous to these peptides.

[0183] Where the peptide is not secreted into the medium, which is typically the case with enzymes, the protein can be isolated from the host cell by standard disruption procedures, including freeze thaw, sonication, mechanical disruption, use of lysing agents and the like. The peptide can then be recovered and purified by well-known purification methods including ammonium sulfate precipitation, acid extraction, anion or cationic exchange chromatography, phosphocellulose chromatography, hydrophobic-interaction chromatography, affinity chromatography, hydroxylapatite chromatography, lectin chromatography, or high performance liquid chromatography.

[0184] It is also understood that depending upon the host cell in recombinant production of the peptides described herein, the peptides can have various glycosylation patterns, depending upon the cell, or maybe non-glycosylated as when produced in bacteria. In addition, the peptides may include an initial modified methionine in some cases as a result of a host-mediated process.

[0185] Uses of Vectors and Host Cells

[0186] The recombinant host cells expressing the peptides described herein have a variety of uses. First, the cells are useful for producing a enzyme protein or peptide that can be further purified to produce desired amounts of enzyme protein or fragments. Thus, host cells containing expression vectors are useful for peptide production.

[0187] Host cells are also useful for conducting cell-based assays involving the enzyme protein or enzyme protein fragments, such as those described above as well as other formats known in the art. Thus, a recombinant host cell expressing a native enzyme protein is useful for assaying compounds that stimulate or inhibit enzyme protein function.

[0188] Host cells are also useful for identifying enzyme protein mutants in which these functions are affected. If the mutants naturally occur and give rise to a pathology, host cells containing the mutations are useful to assay compounds that have a desired effect on the mutant enzyme protein (for example, stimulating or inhibiting function) which may not be indicated by their effect on the native enzyme protein.

[0189] Genetically engineered host cells can be further used to produce non-human transgenic animals. A transgenic animal is preferably a mammal, for example a rodent, such as a rat or mouse, in which one or more of the cells of the animal include a transgene. A transgene is exogenous DNA which is integrated into the genome of a cell from which a transgenic animal develops and which remains in the genome of the mature animal in one or more cell types or tissues of the transgenic animal. These animals are useful for studying the function of a enzyme protein and identifying and evaluating modulators of enzyme protein activity. Other examples of transgenic animals include non-human primates, sheep, dogs, cows, goats, chickens, and amphibians.

[0190] A transgenic animal can be produced by introducing nucleic acid into the male pronuclei of a fertilized oocyte, e.g., by microinjection, retroviral infection, and allowing the oocyte to develop in a pseudopregnant female foster animal. Any of the enzyme protein nucleotide sequences can be introduced as a transgene into the genome of a non-human animal, such as a mouse.

[0191] Any of the regulatory or other sequences useful in expression vectors can form part of the transgenic sequence. This includes intronic sequences and polyadenylation signals, if not already included. A tissue-specific regulatory sequence(s) can be operably linked to the transgene to direct expression of the enzyme protein to particular cells.

[0192] Methods for generating transgenic animals via embryo manipulation and microinjection, particularly animals such as mice, have become conventional in the art and are described, for example, in U.S. Pat. Nos. 4,736,866 and 4,870,009, both by Leder et al., U.S. Pat. No. 4,873,191 by Wagner et al. and in Hogan, B., Manipulating the Mouse Embryo, (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1986). Similar methods are used for production of other transgenic animals. A transgenic founder animal can be identified based upon the presence of the transgene in its genome and/or expression of transgenic mRNA in tissues or cells of the animals. A transgenic founder animal can then be used to breed additional animals carrying the transgene. Moreover, transgenic animals carrying a transgene can further be bred to other transgenic animals carrying other transgenes. A transgenic animal also includes animals in which the entire animal or tissues in the animal have been produced using the homologously recombinant host cells described herein.

[0193] In another embodiment, transgenic non-human animals can be produced which contain selected systems that allow for regulated expression of the transgene. One example of such a system is the cre/loxP recombinase system of bacteriophage P1. For a description of the cre/loxP recombinase system, see, e.g., Lakso et al. PNAS 89:6232-6236 (1992). Another example of a recombinase system is the FLP recombinase system of S. cerevisiae (O'Gorman et al. Science 251:1351-1355 (1991). If a cre/loxP recombinase system is used to regulate expression of the transgene, animals containing transgenes encoding both the Cre recombinase and a selected protein is required. Such animals can be provided through the construction of “double” transgenic animals, e.g., by mating two transgenic animals, one containing a transgene encoding a selected protein and the other containing a transgene encoding a recombinase.

[0194] Clones of the non-human transgenic animals described herein can also be produced according to the methods described in Wilmut, I. et al. Nature 385:810-813 (1997) and PCT International Publication Nos. WO 97/07668 and WO 97/07669. In brief, a cell, e.g., a somatic cell, from the transgenic animal can be isolated and induced to exit the growth cycle and enter G_(o) phase. The quiescent cell can then be fused, e.g., through the use of electrical pulses, to an enucleated oocyte from an animal of the same species from which the quiescent cell is isolated. The reconstructed oocyte is then cultured such that it develops to morula or blastocyst and then transferred to pseudopregnant female foster animal. The offspring born of this female foster animal will be a clone of the animal from which the cell, e.g., the somatic cell, is isolated.

[0195] Transgenic animals containing recombinant cells that express the peptides described herein are useful to conduct the assays described herein in an in vivo context. Accordingly, the various physiological factors that are present in vivo and that could effect substrate-binding, enzyme protein activation, and signal transduction, may not be evident from in vitro cell-free or cell-based assays. Accordingly, it is useful to provide non-human transgenic animals to assay in vivo enzyme protein function, including substrate interaction, the effect of specific mutant enzyme proteins on enzyme protein function and substrate interaction, and the effect of chimeric enzyme proteins. It is also possible to assess the effect of null mutations, that is, mutations that substantially or completely eliminate one or more enzyme protein functions.

[0196] All publications and patents mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described method and system of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the above-described modes for carrying out the invention which are obvious to those skilled in the field of molecular biology or related fields are intended to be within the scope of the following claims.

1 4 1 4239 DNA Homo sapiens 1 tggcagtggg cggcgtagag cactgcagca gcaatgacgg agggcacgtg tctgcggcgc 60 cgagggggcc cctacaagac cgagcccgcc accgacctcg gccgctggcg actcaactgc 120 gagaggggcc ggcagacgtg gacctacctg caggacgagc gcgccggccg cgagcagacc 180 ggcctggaag cctacgccct ggggctggac accaagaatt actttaagga cttgcccaaa 240 gcccacaccg cctttgaggg ggctctgaac gggatgacat tttacgtggg gctgcaggct 300 gaggatgggc actggacggg tgattatggt ggcccacttt tcctcctgcc aggcctcctg 360 atcacttgcc acgtggcacg catccctctg ccagccggat acagagaaga gattgtgcgg 420 tacctgcggc acattgagga taagtccacc gtgtttggga ctgcgctcaa ctatgtgtct 480 ctcagaattc tgggtgttgg gcctgacgat cctgacctgg tacgagcccg gaacattctt 540 cacaagaaag gtggtgctgt ggccatcccc tcctggggga agttctggct ggctgtcctg 600 aatgtttaca gctgggaagg cctcaatacc ctgttcccag agatgtggct gtttcctgac 660 tgggcaccgg cacacccctc cacactctgg tgccactgcc ggcaggtgta cctgcccatg 720 agctactgct acgccgttcg gctgagtgcc gcggaagacc cgctggtcca gagcctccgc 780 caggagctct atgtggagga cttcgccagc attgactggc tggcgcagag gaacaacgtg 840 gcccccgacg agctgtacac gccgcacagc tggctgctcc gcgtggtata tgcgctcctc 900 aacctgtatg agcaccacca cagtgcccac ctgcggcagc gggccgtgca gaagctgtat 960 gaacacattg tggccgacga ccgattcacc aagagcatca gcatcggccc gatctcgaaa 1020 accatcaaca tgcttgtgcg ctggtatgtg gacgggcccg cctccactgc cttccaggag 1080 catgtctcca gaatcccgga ctatctctgg atgggccttg acggcatgaa aatgcagggc 1140 accaacggct cacagatctg ggacaccgca ttcgccatcc aggctctgct tgaggcgggc 1200 gggcaccaca ggcccgagtt ttcgtcctgc ctgcagaagg ctcatgagtt cctgaggctc 1260 tcacaggtcc cagataaccc tcccgactac cagaagtact accgccagat gcgcaagggt 1320 ggcttctcct tcagtacgct ggactgcggc tggatcgttt ctgactgcac ggctgaggcc 1380 ttgaaggctg tgctgctcct gcaggagaag tgtccccatg tcaccgagca catccccaga 1440 gaacggctct gcgatgctgt ggctgtgctg ctgaacatga gaaatccaga tggagggttc 1500 gccacctatg agaccaagcg tggggggcac ttgctggagc tgctgaaccc ctcggaggtc 1560 ttcggggaca tcatgattga ctacacctat gtggagtgca cctcagccgt gatgcaggcg 1620 cttaagtatt tccacaagcg tttcccggag cacagggcag cggagatccg ggagaccctc 1680 acgcagggct tagagttctg tcggcggcag cagagggccg atggctcctg ggaaggctcc 1740 tggggagttt gcttcaccta cggcacctgg tttggcctgg aggccttcgc ctgtatgggg 1800 cagacctacc gagatgggac tgcctgtgca gaggtctccc gggcctgtga cttcctgctg 1860 tcccggcaga tggcagacgg aggctggggg gaggactttg agtcctgcga ggagcggcgt 1920 tatgtgcaga gtgcccagtc ccagatccac aacacatgct gggccatgat ggggctgatg 1980 gccgttcggc atcctgacat cgaggcccag gagagaggag tccggtgtct acttgagaaa 2040 cagctcccca atggcgactg gccgcaggaa aacattgctg gggtcttcaa caagtcctgt 2100 gccatctcct acacgagcta caggaacatc ttccccatct gggccctcgg ccgcttctcc 2160 cagctgtacc ctgagagagc ccttgctggc cacccctgag aacatgccta cctgctgggt 2220 gccgtctgtg cgttccagtg aggccaaggg gtcctggccg ggttggggag ccctcccata 2280 accctgtctt gggctccaac ccctcaacct ctatctcata gatgtgaatc tgggggccag 2340 gctggaggca gggatgggga cagggtgggt ggcttagact cttgattttt actgtaggtt 2400 catttctgaa agtagcttgt cgggcttggg tgaggaaggg ggcacaggag ccgtgacccc 2460 tgaggaggca cagcgccttc tgccacctct gggcacggcc tcaaggtagt gaggctagga 2520 ggttttttct gaccaatagc tgagttcttg ggagaggagc agctgtgcct gtgtgattcc 2580 ttagtgtcga gtgggctctg ggctggggtc ggccctgggc aggcttctcc tgcacctttt 2640 gtctgctggg ctgagggaca cgagggcaac cctgtgacaa tggcaggtag tgtgcatccg 2700 tgaatagccc agtgcggggg ttgctcatgg agcatcctga ggccgtgcag cagggagccc 2760 catgcccctg ggtcgtgagc ttgcctgcgt atggggtggt gtcatggagc ctcatgcccc 2820 tgggtcgtga gctcgcctga gtatggggtg gtgtcatgga gccgcatacc cctgggttgt 2880 gagctcgcct gcatatgcag ggtctgtcat ggaacatccc aagtctgtgc agcagggagc 2940 cccatgcccc tgggacatga acccacctgc gtggaatgct gtttgtgagg tgtctacagg 3000 gtttatagta gtcttgtgga cacagaaatg cacaggggac acttacggac acagaaatgc 3060 acaggggagg ccgagcataa ccaggggtga ggggcaggca gcagttgtag ttactgccgc 3120 ggggcactgc tatgtgcagg gacagccagc gcccagccca tcaccactcc ctgggctggc 3180 tggcaggtat ggcaccctgg gagcccggca tatacccagg gcacccctac ggctgccgcc 3240 agtctcatgc ccaggtgggt gctctgggct ggagcgaggg ccaggttttg ggccgaggct 3300 tccccaggca atcctgtgag ctcccttcta gcctctgacc cagtctggtc tggcttgcat 3360 ggatgtaggg cttggggtgg gaagttcagg tcctggcttt gcctttgcct gatgtggatg 3420 agcagctcac atgctcaggg ccacctgaga ctgtcactgc tctcccctgg ctactgggag 3480 gagtcactga gagcttcgtt acccctgctg ccttgcccag ggcacaccct atacctcctc 3540 atctgctctt cccctccctg ccgccttctg ggcaggtagc agtccctggc ctctccccct 3600 ggctgatcac tctccctcag gcagtggaga tctgcgtctg gacaccctca gatcctgtca 3660 ttgcctgccc agagtccttc aggggcaccc ctctgccttg gtgtgcggtc cagggctctc 3720 acccaggtgc cgcaccctct ggggtcttct gtccagctcc cttgccccat gtgctgtcac 3780 tgactctcct tgggactcgc ctgcctgctc agagccctgc agggcttggt cagctgcctg 3840 ttcagtgtca acacttccct gcacatctta aaactgggct ttattttcgc tgaaggaact 3900 gtgttgggac ccttgacatc tgtcaggttt gcacatgctg tttttttttc tcagcccacg 3960 tgttctcccc cacgtggggt agcagcagga cagacagtga atcacagagt ctgccctgag 4020 cagaggctgc tgtccctggg actcctagcc atggtcagac tgtacaaaac ggttttccag 4080 aaatgaaatg taaatccatt tttatactga aaatgttact gaaagtcact tttatgagca 4140 tctgccttaa taaacagaca ttgattccct taaaaaaaaa aaaaaaaaaa aaaaaaaaaa 4200 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaa 4239 2 721 PRT Human 2 Met Thr Glu Gly Thr Cys Leu Arg Arg Arg Gly Gly Pro Tyr Lys Thr 1 5 10 15 Glu Pro Ala Thr Asp Leu Gly Arg Trp Arg Leu Asn Cys Glu Arg Gly 20 25 30 Arg Gln Thr Trp Thr Tyr Leu Gln Asp Glu Arg Ala Gly Arg Glu Gln 35 40 45 Thr Gly Leu Glu Ala Tyr Ala Leu Gly Leu Asp Thr Lys Asn Tyr Phe 50 55 60 Lys Asp Leu Pro Lys Ala His Thr Ala Phe Glu Gly Ala Leu Asn Gly 65 70 75 80 Met Thr Phe Tyr Val Gly Leu Gln Ala Glu Asp Gly His Trp Thr Gly 85 90 95 Asp Tyr Gly Gly Pro Leu Phe Leu Leu Pro Gly Leu Leu Ile Thr Cys 100 105 110 His Val Ala Arg Ile Pro Leu Pro Ala Gly Tyr Arg Glu Glu Ile Val 115 120 125 Arg Tyr Leu Arg His Ile Glu Asp Lys Ser Thr Val Phe Gly Thr Ala 130 135 140 Leu Asn Tyr Val Ser Leu Arg Ile Leu Gly Val Gly Pro Asp Asp Pro 145 150 155 160 Asp Leu Val Arg Ala Arg Asn Ile Leu His Lys Lys Gly Gly Ala Val 165 170 175 Ala Ile Pro Ser Trp Gly Lys Phe Trp Leu Ala Val Leu Asn Val Tyr 180 185 190 Ser Trp Glu Gly Leu Asn Thr Leu Phe Pro Glu Met Trp Leu Phe Pro 195 200 205 Asp Trp Ala Pro Ala His Pro Ser Thr Leu Trp Cys His Cys Arg Gln 210 215 220 Val Tyr Leu Pro Met Ser Tyr Cys Tyr Ala Val Arg Leu Ser Ala Ala 225 230 235 240 Glu Asp Pro Leu Val Gln Ser Leu Arg Gln Glu Leu Tyr Val Glu Asp 245 250 255 Phe Ala Ser Ile Asp Trp Leu Ala Gln Arg Asn Asn Val Ala Pro Asp 260 265 270 Glu Leu Tyr Thr Pro His Ser Trp Leu Leu Arg Val Val Tyr Ala Leu 275 280 285 Leu Asn Leu Tyr Glu His His His Ser Ala His Leu Arg Gln Arg Ala 290 295 300 Val Gln Lys Leu Tyr Glu His Ile Val Ala Asp Asp Arg Phe Thr Lys 305 310 315 320 Ser Ile Ser Ile Gly Pro Ile Ser Lys Thr Ile Asn Met Leu Val Arg 325 330 335 Trp Tyr Val Asp Gly Pro Ala Ser Thr Ala Phe Gln Glu His Val Ser 340 345 350 Arg Ile Pro Asp Tyr Leu Trp Met Gly Leu Asp Gly Met Lys Met Gln 355 360 365 Gly Thr Asn Gly Ser Gln Ile Trp Asp Thr Ala Phe Ala Ile Gln Ala 370 375 380 Leu Leu Glu Ala Gly Gly His His Arg Pro Glu Phe Ser Ser Cys Leu 385 390 395 400 Gln Lys Ala His Glu Phe Leu Arg Leu Ser Gln Val Pro Asp Asn Pro 405 410 415 Pro Asp Tyr Gln Lys Tyr Tyr Arg Gln Met Arg Lys Gly Gly Phe Ser 420 425 430 Phe Ser Thr Leu Asp Cys Gly Trp Ile Val Ser Asp Cys Thr Ala Glu 435 440 445 Ala Leu Lys Ala Val Leu Leu Leu Gln Glu Lys Cys Pro His Val Thr 450 455 460 Glu His Ile Pro Arg Glu Arg Leu Cys Asp Ala Val Ala Val Leu Leu 465 470 475 480 Asn Met Arg Asn Pro Asp Gly Gly Phe Ala Thr Tyr Glu Thr Lys Arg 485 490 495 Gly Gly His Leu Leu Glu Leu Leu Asn Pro Ser Glu Val Phe Gly Asp 500 505 510 Ile Met Ile Asp Tyr Thr Tyr Val Glu Cys Thr Ser Ala Val Met Gln 515 520 525 Ala Leu Lys Tyr Phe His Lys Arg Phe Pro Glu His Arg Ala Ala Glu 530 535 540 Ile Arg Glu Thr Leu Thr Gln Gly Leu Glu Phe Cys Arg Arg Gln Gln 545 550 555 560 Arg Ala Asp Gly Ser Trp Glu Gly Ser Trp Gly Val Cys Phe Thr Tyr 565 570 575 Gly Thr Trp Phe Gly Leu Glu Ala Phe Ala Cys Met Gly Gln Thr Tyr 580 585 590 Arg Asp Gly Thr Ala Cys Ala Glu Val Ser Arg Ala Cys Asp Phe Leu 595 600 605 Leu Ser Arg Gln Met Ala Asp Gly Gly Trp Gly Glu Asp Phe Glu Ser 610 615 620 Cys Glu Glu Arg Arg Tyr Val Gln Ser Ala Gln Ser Gln Ile His Asn 625 630 635 640 Thr Cys Trp Ala Met Met Gly Leu Met Ala Val Arg His Pro Asp Ile 645 650 655 Glu Ala Gln Glu Arg Gly Val Arg Cys Leu Leu Glu Lys Gln Leu Pro 660 665 670 Asn Gly Asp Trp Pro Gln Glu Asn Ile Ala Gly Val Phe Asn Lys Ser 675 680 685 Cys Ala Ile Ser Tyr Thr Ser Tyr Arg Asn Ile Phe Pro Ile Trp Ala 690 695 700 Leu Gly Arg Phe Ser Gln Leu Tyr Pro Glu Arg Ala Leu Ala Gly His 705 710 715 720 Pro 3 42450 DNA Homo sapiens 3 tcatgactgc ccctagaagc ttaactgtgt caattctcag acgtagttta cagctttttc 60 ttttctttca gacattaaaa agagcggatt attttactca taaaaagtcc agtccattaa 120 gatatcaaaa ctcaaactct tatccagttg aaacctcttc cctcacctag ctttgccagg 180 ttcagtgtga gattccatcc aggctgaagc cccttatccc tattcttcat gtttctacat 240 ggaggaactt acctggagaa aaacttccag cctctttctg cttccagaga agtagagtga 300 ctcatttgat tgaatttcag agaacagata gggtggagtg tgctcaggct cctctgggta 360 ctctttctgg ggtctgtggg ttgactggag gggtgtcttc tggtgggcac tcaattgcat 420 agtgcttggt gaggcagttt catggcctag aggctggggg atatgtttgt ctgacttacg 480 ggtgatttag tagcttgccc tcttgcttgc agatttaagc cttgtccttc aagctaggtt 540 tttaatttgt ggcaaagctg atattttgat acccacccat cttattgctg tgtctttttc 600 atccgtttct gaactgggat aggaagaggt gattatcctt gattgtctaa aaccccgcta 660 ttccactgtg gggaaggtgc ctgtgggtat tcttttgtcc actctctctt ccaactttct 720 cctccggctt gctgtggctc accgcccctt cgaagttagg ctgggggtag gaattgagga 780 gtgggtgccg aaatgctcac taggctgggg cagttgtaac tggatgtcag ggcttctgtg 840 ggccaggtga agacatgctg gggtcttctg tgggtccttg acctgactta gggaccactg 900 gctgcagcct ccagacgtca gccatgtttc caacagtcag acgccccctg ccctgttgcg 960 cccggctgtc ccttccaagt tcggtcactc gctctgcctc catcttcctc ttccctctgc 1020 tgctaaggct tttcaccttt aatttctcct ggggccaccc ccaactccag cgaccccgtg 1080 agcagctgag gctctaccgc gctcggtcct ggccagcgac gcagcccttc cctggcgggg 1140 ctccagggct tctggcccct gtggtccgcc aggtgtgggg gcccacggcc tcaccgcgcc 1200 taccccactc cccccggcga agctacgcgg cgctcagctt cccagggacg ccggcggcgc 1260 cctcggctcc tccgctccgc cccgccctcc ccctggtctc gcactggagc cgacggcccg 1320 cgcccacctc acctcagggc ggcctcccgc ccccaccccc ggccccggcg tccgggcaaa 1380 tcctgcagcg cgagagcaat tccctgccac ccgaccttcg cactcgctgt cgctcgctcg 1440 agcctcgctc cccacgtcct tccttccgac ccgcggctgg accctcctca caaatttctc 1500 agagaggctc acctcaaagc gcggcgcacg aggccgggct cccgggacgc aagcctctag 1560 agggcgcgcg agaggccccg cccccgccct tcggccccac ccaccagccc cgcccccacc 1620 cgcacccacc aggccccgcc cccacctccc cacccaccag ccccgccccc acctccccac 1680 ccaccagccc cgcccctcat gccccgccaa taaggcccca cccgcctccc ccgtcccgtc 1740 gccttcaccc accatccccg ctccctcagg ccccgcccca cgccgcatgg ggcaccaagc 1800 gctccaccac tgtggtcgcc tggcacaccc cggggtcacg ctcgcggcgc tctgattggt 1860 tgcgtgggcg tcggcccacc taagcctgag cgcctgccga ggcctgcgcc tgcgtagtgc 1920 gcgcgggagg ggcgggaggg gcgggagggg cgggaggggc ggggctgggc ggcaggtccc 1980 gggtgcggac atctggcagc tggcagtggg cggcgtagag cactgcagca gcaatgacgg 2040 agggcacgtg agtcccctcg ccccgggctc ctgacgaatg cggggtggtc ctaggtgctg 2100 aggagagcgc gactggggca gtgggccggc ggccggcgtt ggggcggggc ctgggtcgct 2160 gatggccggt ggtcctcagg tgtctgcggc gccgaggggg cccctacaag accgagcccg 2220 ccaccgacct cggccgctgg cgactcaact gcgagagggg ccggcagacg tggacctacc 2280 tgcaggacga gcgcgccggc cgcgagcaga ccggcctgga agcctacgcc ctggggctgg 2340 acaccgtaag ttgcttccgc ggagcgtcag cgagctcggg accctgaggg gtgagccgtg 2400 aggagcacgt tttctctcag aaaggcgggt gggaggaccc ggccagcgac gcccatcccc 2460 aaggcgagcg cccacgggaa ctgcgttcgc gggcccctcc gcttcagccc cttcatctct 2520 aaaccacgca taggagactc ctaatgtttt attttttagc accttatttt gagataattt 2580 ttgacttata ggagagttgc aaagatagtt gtaactttgt ttttattcac aaaaagtgtt 2640 tggatccact gtcttagttg tgtgcattgt aagagatttt ggtcgtcaga gtctgcagtg 2700 taaacagggt ctcctgccga gccccggcca ccgagggaaa ggctgtgccg ccccttgggc 2760 cctctttgag aggcccgagt cccaggccca ggtcggcacc cgtgccccac cctacagtct 2820 gggtgcctgg tttattccag acatcttgga gaagttgtga agaatacatg actggcaaat 2880 aaagcaacga aaatgtgcag ctgttctttt actttgctga ggtgtgatgc tctcatcaaa 2940 gagtttcaga cttttgatgg aaacagctga aacttttaaa gtaatttaca ttcactgttt 3000 tgacttgggc tgtatgtgaa gagggttcct ctggccgggc aacagtcccg tcagctatct 3060 cttttttttt ttttcgatct ctttgcagaa gaattacttt aaggacttgc ccaaagccca 3120 caccgccttt gagggggctc tgaacgggat gacattttac gtggggctgc aggctgagga 3180 tgggcactgg acgggtgatt atggtggccc acttttcctc ctgccaggta ggagtatgct 3240 gccccagcct gatggtatgg ccaccctgga tcacccttgg gatcctggcc cagcctggtc 3300 tagggttttg atgaagcagg tgaaaatcca ggggctcaca agaaaagggc tggcaaactc 3360 tgccctatgt cagagtcgtc ctgctattgg tctaggggat cagctagcct tgccagtgta 3420 gggtgacagg ctctctgata agagaagcaa gtggttctct agggctctgt gttgccttga 3480 gggaggagga aggtgggctt tgaagtctca gtacaggatg ggatggacat tccaggtgga 3540 aggcccagcc tatgccaagg ggctgtaggt gggcagagtg gtgggtgggg agctgatatc 3600 tgctgtgaac ttcctcgggg ctattgcagg agagcttcag gttcaggctg gtgagtagga 3660 ggagcatagc agttggactg cctgggtatt gaactgattt ggctacacaa gactattttg 3720 catcctggga gtgtttctct acagaaatcc tcagccttgt aaaatgggaa attccctcct 3780 atgaatttat gcaataggac ttttttccct agtgacttgt aatcacattg tttcaatgac 3840 gtgaattcct acataaatag gttttgtttc tgtgataact cttactgata catcattttc 3900 ttttactacg ctgactttgt aatagataga aagtccttat atacctttgt tgcctttctt 3960 tttaaaacat ctcttacctg tgtctattca tttactcatc caaattgcct ttatcctgat 4020 tttgtcccag acttgaaatg aagttgcaat aggcttatat gttagtttgg gaagagttgg 4080 cctttaacgt taaaaacagt tccatggtgt ttactgtagg ccaagccctg ctcaaggcct 4140 gttcttcttt tagtccttag aataagccta atgagataca ttagaaagct gaggcacatt 4200 tattccaggt aaccagacta gcaggaggag cactgggatc cccatctctg ctttgacttc 4260 tagccctgct gccacctgga ctgtacagca ttgagttttt ctgtcctggg atttgagggc 4320 ctgtccttag gggaagtcaa ggtgctcttc ttcccttggc cccatcaggg cctgtttaga 4380 ctgttctcag ggctcgtggt aaggcaatga catagagttg gtcaggagat gggtcagccc 4440 cactttgcct ctgtagcctg acctgtgaca ggattggaat caggtttggt catgtgcaca 4500 gtgtcaggca tgcagtggtg cttggtcagt ggggattact gtgttgtttg ttcttgctgc 4560 tttggctctg ggcttagctg gctgggaccc ttcctgtggg ctggctgtga gttggagttt 4620 ttttgtattt tttttttttt tttgagacag cgttcgctct tgttccccag gctggagtgc 4680 aatggcacaa ttttggctcg ttgcagcctc tgcctcctgg gtgcaagtga ttctcctgcc 4740 tcagcctcct gtagggtcca gccccacagg gtcggtaggt ttttctccct gtgtgcggag 4800 atgagagatt gtagaaataa agacacaaga cagagagatg aaagaaaaga cagctgggcc 4860 ccgggggacc actaccacca agacgtggaa accggtagtg gccctgaatg ccaggctgcg 4920 ctgatattta ttggatacaa gacaaagggg cagggtaagg agtgtgagcc atctccaatg 4980 ataggtaagg tcacatgggt cacgtgtcca ctggacagtg ggcccttccc tgcctggcag 5040 ccgaggcaga gagtgggaga gagagagaga gagacagctt atgccattat ttctgcatat 5100 cagagacttt tagtactttc actaattttg ctactgttat ctaaaaggca gagccaggtg 5160 tacagggtgg aacatgaaag tggactagga gcgtgaccac tgaagcacag catcacaggg 5220 agatggttag gcctccggat aactgcgggt gggcctgact gatgtcaggc cgtcccacaa 5280 gaggtggagg agtagagtct tctctaaact cccccgggga aagggagatt ccctttcccg 5340 gtatgctaag tagcgggtgt ttttccttga cactgacgct accgctagac cacggttggg 5400 tccgcttggc aacgggcctc ttcccagatg ctggcgttac cgctagacca aggagccctc 5460 tagtggcctt gtccgggctt aacagaaggc tctcactctt gtcttctggt cacttctcac 5520 tatgtctctt cagctcctat ctctgtatgg cctggttttt cctaggttat gattgtagag 5580 cgaggattat tataatattg gaataaagag taattgctac aaactaatga ttaatgatat 5640 tcatatataa tcatatgtat gatctagatc tagtataact cttgttgttt tatatatttt 5700 attatactgg aacagctcgt gccctcggtc tcttgccttg gcaccaaggt ggcttgccac 5760 ccacagcctc tcgagtagct gggattacag ccatgtgcca ccatgcctgg ctaatttttg 5820 tatttttggt agagacaggt tttcaccttg ttggtcaggc tggtctcgaa ctcctgacct 5880 cgtgatcccc caccccccac ccccagcctc ccaaagtgct gggattacag gcgtgagcca 5940 ctgcacctgg ctgagttgga gcttttcttc cctctttttg gactttggaa aatgctcttg 6000 gtccatgatg ctatgtagac agctcccgtt gactgtggcc tgtgcggcat tgggcagcac 6060 tctggtgaac actgaatcgg gtctgacctc ctagccccac catttactgg ctgagcctca 6120 gtttccttgc ctgtaaaatc aggaagatgc tggctctgct cctctctgca catttccccg 6180 tcctaacaac attataactg ttaggaaaga gacgggcttg ttttgggatg gctcatttta 6240 tgtgaccctg tgcgctgtct ctgagtccat ctgcccttct tccagggtgt agggaccagc 6300 cccacagggt cggtgggtct ctccctgtgt gcggcgatga gagagtgtag aaataaagac 6360 acaagacaaa gagataaaag acagctgggc ccgggggacc actgccacca atgcatggag 6420 accagtagtg gccccgaatg tctggctgtg ctgttattta ttggatacaa agcaaaaggg 6480 gcagggtaaa gagtgtgagt catctccagt gataggtaag gtcacatggg tcacgtgtcc 6540 actgggacag ggggcccttc cctgcctggc agccgaggca gagagaggag acacagagaa 6600 agaaaactta tgccattatt tctgcatatc agagactttt agtactttca ctaattgact 6660 actgctatct agaaggcaga gccaggtgta caggatggaa catgaaggcg gactaggagc 6720 gtgaccactg aagcacagca tcacagggag acaggcctcc ggataactgc gggcaggtct 6780 gactaatgtg aggccctcca caagaggtgg aggagcagag tcttctctaa attcccccgg 6840 ggaaagggag cctccctttc ccggtctgct aagtagcggg tgttgttcct tgacactttt 6900 cgctaccgct agaccaccgt ccgctcggca acgggcgtct tcccagacgc tggcgttacc 6960 actagaccaa ggagcccttt tgctggcccc gtccgggcat aacagaaggc tcgcactcct 7020 gtcttctggt cacacctcac tatgtcccct cagctcctat ctctgtatgg cctggttttt 7080 cctaggttat gattgtagag cgaggattat tataatattg ggataaagag taattactac 7140 aaactaatga ttaatgatat tcatatatct ctaagatcta tatctggtat aactattctt 7200 gttttatatt ttattatact ggaacagctc gtgtcctcgg tctcttgcct tggcgcctgg 7260 gtggcttgcc gcccacacag ggcatgtctg gatggtttga acactagggc ttctgatgct 7320 ctaagccaga gtcaggtatt cattccatgg cacatgtggc tggggtctgc cctgagacct 7380 gtcccgtgcc aggctctggg ggcacatggc tgatggaacc aagcatgggg agtgaaggtg 7440 gagggtggcc tgtgagcacc atgcctgaga ggaccaggct ggggacggaa ggttcttagt 7500 ggataatatt tattgtctct gcctcccccc tgacatttgc aaagcggcat atgcttgtaa 7560 aaaaattttg aaacagaaaa atataaataa ataagtaggt attaccacat gcaagggtga 7620 ccaattttgt atttttcttc ccagcagatg ttaaagcaag accaacagtc tcccctcatg 7680 gaaggcccac tgatctaaaa tgctggttcc ttttggacct tcagggcact tgggggagac 7740 cttcctgagg tgctgtgcag tgtctggtgt ttctcagacc caggtggtca tgggagccag 7800 gcgtggctga gtgggctcta caggccctag gcagggagca tcgcctgtgc tgtggctgac 7860 gttccttctg gccctgttcc caaagttccc catgggggcc tgggaggaat ggcctttcca 7920 gggggtgttt ttatgagaag gaggtagctc cctgttggag tgaggtgctc aggaggaaag 7980 gggcctggtc ttagcagtca tgaccacctg tccccagtga ggaacatctc tcctgccaca 8040 caggcctcct gatcacttgc cacgtggcac gcatccctct gccagccgga tacagagaag 8100 agattgtgcg gtacctgcgg tcagtgcagc tccctgacgg tggctggggc ctgtgagtgt 8160 gcctgcccct gtgtcactgc acatgtgcat gtgtgtgttc tcatgatgta ggagatgctt 8220 gggtttccag gcagctgcca ggggttagga gtgattgcag ctgtgggtgt ggggtgggtg 8280 agggagagac tagcaggcgg ggagtgggct gaaggccatg caggtggggc ctcggcttca 8340 catcttttgt taaatggatt ttgtggctgt tacgacactc ttgagaccca catgtgaaaa 8400 ctgtcagtct gttatcactt aagacagaag aaaattgccc ttgactctgg gctggcagca 8460 ggtggagaca aggcctgaca gctttcctgc catgtggcac acactttggg agcagagcca 8520 tagcccaaag tggaccgccc ttgagctaga agtgttgact caggcgtggg aaggtgtaga 8580 gcaggcgggt cacggtgagg aaggagtggg gggctcagtt gtcatgggag gtgcatgaat 8640 tcgtactgca gagtggctgc tcaggggtct cctgtgttga catgttatgt caggttaagc 8700 cattttagca ttcttagttt tctgaggaaa ctccacagaa agttttgctt tatttcttag 8760 aagtaaggac agataccggt ttctcacctg tcctctgctc ctgtaggcac attgaggata 8820 agtccaccgt gtttgggact gcgctcaact atgtgtctct cagaattctg ggtgttgggc 8880 ctgacgatcc tgacctggta cgagcccgga acattcttca caagaaaggt acggcatgtg 8940 cagcatgtgc tgggccaggg gttcgtgtca actcgataat gagctctcac aaacgagata 9000 cagaaagatg cacttgcagc tgaaacagtg ggcaaaagca catgagcagg gaatttgtca 9060 aagcagaagt aggcagacac tgtttaacct aggcatcatt ttttaaaaaa gcaaattaag 9120 agccaggcac agtgagtggc tcacgcctgc aattccagca ctttgggaga ctgaggtaga 9180 aggaccactt caacctaaga gttcgaggcc agcctgggca acatagtgag acctggtctc 9240 tacaaaaaca ataaaatatt agccaggtgt gatgatatgc acctgtagtc tcagctactt 9300 ggaggctagt aaggcaggag gatcacttga gcccaggagt tctgggttgc aatgagctgg 9360 ttgtactact gcactctagc ctgggtgaca gagtgcgacc ctgtctctaa taaaataaaa 9420 aagccaagca aactaagaca accaggtaat tctgtttgtt tcctgaattg gcaaaaactt 9480 aaacgaaccg tgttaatatg tccaccttct ggggggcagc ctggctgcag gcaagagcag 9540 ccctggagct tgcaccttcc aagctgatcg tctacctctc caagcccggg gctgtccacc 9600 tctccaagcc cggggctgtc cacctctcca agcccggggc tgtccacctc tccaagcccc 9660 gggctgtcca cctctccaag ccccgggttg tcttacctct ccaagcccca actgtctact 9720 tctccaagcc ctggtctggc tacctctcca agccctgggc tgtccacctc tccaagcgcc 9780 aactatcttt ctctccaagc cctggcctgg ctacctctcc aagccccagg ctgtccacct 9840 ctccaagccc caactgtcta cctctccaag ccccggcctg gctacctctc caagcccctg 9900 gctacctctc catgcccggc ctggctacct ctcctcttgc ctataggccc tgaggggcaa 9960 ttccagccca agggaatcca tggctcctgc tgctccaaga aaacctagtt tatgttgtgg 10020 ctctgcagag cctggcctgg tcttgtcctc tgtgtttcac agaccttccg tagccagtcc 10080 cacctgccct gctctctgct gcatgcgcag gggcctcctg tcagctcctc agagaccctt 10140 attatcccag ggctcgccat gcactgcctc cttcgcctgg agcctcttac cttccactcc 10200 tgccccgctg gctcacactt tacgtgttcc ttctttgagg acctcttcct gacctaccgt 10260 gccaggtgga gtgtcctgtt acgcattctc atgagatcct gccttctttc ttggtgagct 10320 tgtcactatt gtcctcagtt cactgtcagc ctttggtgtc gttgatgctg cgtccccaag 10380 gctgctgtcc ggttcccacc acactcctgg cgcctgcctg gtgaaggaac gtgtttaggc 10440 tgcactttgc ctagtagctt tgtgggtctt tattgacttt tgcatacctt ttggggtttg 10500 gagcagggac tcctcagaag catgtttaga tggtgtggct gtgccaggac tgctgctgct 10560 gaagtggctc tggcatgggg ccagcgtgct ggagctactc tggagtctag ggtcgtcttt 10620 gttcccatac aggaccagtc tgccaagtgg agatgacaca gactggggca gctcaggctt 10680 ggctcagagg gcgaggctga gtgtgcgctg tcacttcccc accttgcctt ctccaggcgc 10740 atgtgcacct gggcccctcg ctcacctgag cactgaggtg tccctggacc ttcccaggta 10800 gctgtcttca tgtgctcctt cctggggcca ggggttgcaa acacctctcc tggggctgga 10860 cacacacact cccaggaaag ccactggttc cacctagggg gccgtgtatc caggcaagtt 10920 ctcagcactc tggaacctgc ttcgcacatg ggggtcgcaa gatccacatg aggctgccct 10980 tgcctcatgg agaggggcac acgtgactcc cagagggtga agcttcccag ctagaggcag 11040 tgcagacttt gctgacagga agcagatgac gtgggcctat tctctccccg ctcaggtggt 11100 gctgtggcca tcccctcctg ggggaagttc tggctggctg tcctgaatgt ttacagctgg 11160 gaaggcctca ataccctgtt cccagagatg tggtatgtct gctgttgatt gggttgttgg 11220 gtcgctgctg ctgtcccggg gagtagagtg acagggaccg tgggtcaggt gcaggctgtg 11280 acagcagaga ggggtgggca ttctgtgggt gggtggagtt aggctcctgg cagaggccct 11340 gatcaagctt gagtcctgta ggggtacaga aagggggagg ttcccaattg agcaggaaga 11400 aggctgtgcc atggatggag gtaccccgag tcaggctgca ggcagggctg ggtggcttcc 11460 ctcttgctgt ggaagactca gcatctgtag aagtgggggg gtgcccctcc cccagcctgc 11520 acaggggcgt cctgtgttgc tgctgctgcg tttgtctcct ttgctggtga atgtgaagtg 11580 tgtcccgacg tgacacctca cctgtggact cagcgtgtgt gcctttaaaa gatcagtgtc 11640 tgtggccagg tggggtggct catgcctgta atcccagcac ttcgggaggc cgaggcgggc 11700 agatcacgag gtcaagggat cgagaccatc ctggccaaca tagtgaaatc ccgtctctac 11760 taaaaataca aaaattagct gggcgtggcg gcgcgtgcct ctagtttcca gctactcggg 11820 aggctgaggc aggagaatca cttgaccctg ggaggcagag gttaccgtga gccgagatcg 11880 tgccaccata ttccagcctg gcgacggagt gagactctgt ctcaaaaaaa aaaaaaaaaa 11940 gatcagtgtt tgttttttta aacagaacca catactgttt aaatacccag caaaatcaac 12000 attaatttct tattatctgg tgtgtgtttt ttttgttttg ttttgagacg gagtctagct 12060 ttgtcactca ggctggagtg cagtggcgtg atttggggtc actgcaacct ccgcctcccg 12120 gattcaagca attctcctgc ctctgcctcc cgagtagctg ggattacagt ctcaggccat 12180 cacgcccagc taattgttgt atttttagta gagacagggt ttcactatgt tggccaggat 12240 ggtctcaaac tcctgacctc aggtgatccg cctgccttgg cctcccaaaa gtgctgggag 12300 ccatgagcca ctgctcccgg ccttatgtgg tgtctttaac cagtgtcttg taacatttta 12360 tggctatcta ttgaaagcag tggacatctc cccagaaaac actcgtgcat atgagtttac 12420 cccgttatgc attttgggaa gtgagaccct ggaaccacac agagcccctg ctggcttcct 12480 tgagtgttgt gggaaccctg gtgggggtgt cccctacaga gctatcatca gggctggggg 12540 ggtcccttgt gttagatgac tttggtgcgg gggtgggggg tggggggtca agttagggga 12600 ggcaggaagt gaaggggccg ctcaagaaag gacagcagca gtgtcctgat gcaaaggccg 12660 ggggcttaac cccggaagcc agtttgggtg gtgacgggga ggcacaggga tggtgagatc 12720 accccgggag ggtagacaga gataccagag tagggggcag ggttagggtg ccgctacctg 12780 aggcgggccg tagagcacat aggttgggag gtgtcctggg gccattcaaa tgcccgctgg 12840 actctgcgcc tcgcccgtgt gtaatgagcg gcagaggaag gactgagacg gcagtcagca 12900 cagctgccag ggcaggaggg gtgtgggttc cacacgctgg tgctggtgag ggcgtctcat 12960 ctgccccact tgggggggcc gtcggtcagt gctgccgcat gggcacgcca gggtgctgct 13020 tgtctttgct ggagttgctt ggagggtggg ttgggaggtg aaaggaggac cacagacctg 13080 aaccactcca gctgcgaaat gctggaagtg taacccaaaa tgtgagaaaa aaaaacaccc 13140 ttttaagtaa gtgggtgtga aagtgggcca aggcctgatg ccacagtcag ggagcaggga 13200 aggctcagca ttgctcaccc tcacttaagg atggggctag catcacataa ggcatcacat 13260 aaggatgggg gctagcaggg aaagggagag aaaacacatg aggcacacac agaccctggg 13320 aagctggtgg agctgtgcta acgtcagcag accagtgatc aaagacccag gccttgggga 13380 gattccacag acctacagac ctacagtttc ttttttcttt ctttttcttt tttttttttt 13440 ttttgagaca gagtctctcg ctctgtcacc aggctgtgtg cagtggcaca atctcggctc 13500 actgcaacct cctcccaggt tcaagcgatt ctcctgcctc agcctcccga gtagctggga 13560 ctagaggcac acaccaccat gcctggctta tttttgtatt tttagtagag atggggtttc 13620 gccatgttgg tcaggctggt ctcaaactcc tgacctcaag tgatccacca gcctcggcct 13680 cccaaagtgc tagggttaca ggcgtgagcc accgtgcccc tcctaaagtt tcttaaatac 13740 acttttaaaa gtaaacttta aattttggag tagtttccaa tttctggaaa agttgcaaag 13800 atagccaaga gtgttccctg gggccctcac accatatccc cattgttgat gttttatgtt 13860 accaaggtac gtttgttgta gctaagaaac ccacggacaa tcctaagcat ttaggagctc 13920 catcacctgg ttttaggatg caaaatgctg accgaactag gaggtgcagc tcctcagagg 13980 gtgcacctat ggttcaactg tgcccctcag gagcacggtt gggaaatgcc cgcagatgca 14040 ctgacgtggt ggggaatagc catccaccag tgttcgtgct tgaaagggcc caaggtatgg 14100 atgctggcgg agggggcagg cttgagtctt ggggtctccc actgactcct gctgttgccc 14160 caggctgttt cctgactggg caccggcaca cccctccaca ctctggtgcc actgccggca 14220 ggtgtacctg cccatgagct actgctacgc cgttcggctg agtgccgcgg aagacccgct 14280 ggtccagagc ctccgccagg taggacctca tcagggaaca aagtgaaggc ctctggggct 14340 gggacccaca gggcctgggg cttctggaat ctaaccacac ctgtccactc acctggtggc 14400 cctgtggagc ggagagccct gtggagcaga gcctccacct tcctccatcc tataataaac 14460 agtgagcaag ctctgcccag aggggacttg tgctatggga cagtcagtag ctgtagccca 14520 gggttcctgg gggggacttc caggactcaa gggatgcagg aggcagatgt gcactgtgtc 14580 ctctggaagc aggcctgagg cgaggtttga ggtgcaggat gtttatcagg cctgccatgg 14640 ggaagaagga ggggcagagg gaggaaatga gcttctgggc agacctggga ctcatggagc 14700 tggggagctc ctcagagcgg tcctcccata gggggccttc atgtgccctc ggggtcagtt 14760 gctggaggga cccccaccca ggaagggact ggcccagggc cctgagggcg gatggtggga 14820 ggccacccct cctggtttga gccaggccta ccaggtgctc ccaggcccca aggctcagac 14880 actgccccta ccaggagctc tatgtggagg acttcgccag cattgactgg ctggcgcaga 14940 ggaacaacgt ggcccccgac gagctgtaca cgccgcacag ctggctgctc cgcgtggtat 15000 atggtgagcg cctcctgagg ggccggcagg gcagcccagg gtcagggtca gggtgtcgcc 15060 cactcattca cgcactcatc ccctgccagc ggcactgggc cacctcctct gtgccaggcc 15120 ccagggggcg ggatctcatc gccctgcccc tccaccctga gaaccagctg gtcttctact 15180 ctcaggagtc caccctgtgc aagggtgtgt ggtaggaggt gtggggcagc ccctcctggg 15240 cagggaagga ggagctcaga gaccaggcct gggggtgggt gggaggggga aaccctgggg 15300 aagggcaagt ccaggcgttg cagtgcatgg agctccaggc tgaggccagt gtcatggtgt 15360 ctggcatcca ctgacccctg tcccctgtag cgctcctcaa cctgtatgag caccaccaca 15420 gtgcccacct gcggcagcgg gccgtgcaga agctgtatga acacattgtg gccgacgacc 15480 gattcaccaa gagcatcagc atcggcccgg tcagtgcccc tgcccggcct ctgactgcag 15540 cccctggggg ttgaggtccg aaagtgaagt cctagaggcc gggctgtgag ctgggagtgg 15600 ggtttcctgg agcctggtgt acctccattt gggaggtggc cctctgatcg cacaagtgtc 15660 tgagggcttc tgtcctggac ccctgcacgc ccagctcagt gaagttgccc caccacactc 15720 gaccccccgc ttccgtcccc caccggctct tgtcctcagt gtgcctggac actctcctag 15780 aggcccctcc ctgagatctt gctggctagc tggctagctg ggaggggtgc tttttcctca 15840 cttggttccc tctccccaaa cagttcatca ttcgccattc tcccgtgggg tttagacatg 15900 cccagggtgg gtgggagtag caggtgccac tcctgattcc tcctgcctag ctagggactt 15960 ggagctctca cctctgtggg gcctgcaggg gtccaggtgt ggccagttca gtgaccttag 16020 agggtgcaat ccccgggctg tgctggtgcg tggccgcctc ctgacagagt cagcaggccc 16080 tgggctgtgc tgcagctgct gccgtagctg tgcgcgtagc tgctgcggtg tagtgggttg 16140 gcttaggcat tctctggaca tacccaggtg gcactgggcc actgagtccc accctgacac 16200 tgcatctcgg attttctggg cctcatgcca cctcagtgga tcacaaatcc tgactgaccc 16260 tgcagcgggt cccttgtttt ttgctcagca gtgatgtggt tctttgtggg ttttggttta 16320 atcccatata gagcacatct gtactaaacg cattagaaac atgcttgcaa ttggatcttg 16380 acttgtgaga tgcataagta aaaagttggg ggcctctgga acattctgtt ctgaggaaga 16440 aggggggcaa gtggtcccta ctgctacagt cctgtcttcg catctcttcc tgggcccctc 16500 aggccctgtc ctctgtcccc tgtgttgtct ctaaggcacc tggtagccca tgcccctctg 16560 gtttctcctg gaacccctcg cttctccctg gtggagtgct gctccttctc acagcctaag 16620 gcaggctgtg gccttggccg acactgcctc tgtctgagtt gggtcctggg gacacagttg 16680 ttgcccatcc tcgctcagga aatgcctgtt agagcagaag gcccctgtcc tggccctgag 16740 tgatctgcac ggcactttat gcctgggggc tgctgtggat ctggacgaga ccttgtccct 16800 ggaggctgct gtgggtctgg agcggagcct tgacagggct gtctctcctg cagatctcga 16860 aaaccatcaa catgcttgtg cgctggtatg tggacgggcc cgcctccact gccttccagg 16920 agcatgtctc cagaatcccg gactatctct ggtgagtgtg gctgggatat gctggcgggg 16980 cctctcacga agactggatc tgagccccag ctgcatccca gtgagggggc ccccacggtg 17040 ccatctggga atactgccag ggaatacctc caggaaccag cagtgtcagg gcttgtggaa 17100 gccactgagg gttgtctttg aattggaaga tttgccaccc agtggaagtg tggggtgttc 17160 ccagaaggta gagtgaggaa gggggtggta ggtagcaggg caggttcagg ttggcatcag 17220 gaggcctgtg gacaagggga gcttgtcagc catggactgt gccctggagg tggggcccct 17280 gtcatggagg gcagagagcc gtcccatggt gggaagcttc cgctgtacag gcctcttcct 17340 ctggtgcctc agcactgcac gagggcggca gggctggcac agcctggggt cggggagcct 17400 cccgctgccc cttgccttgg gtgtggccct tctgggtgag tgtgtcctgt tttccataga 17460 gtgtggccct cacccccagg agcccagcag cccagctggg gtggcatcca ggccagtgcc 17520 aggcctcggg aggggacaga cggcctctct gggaccctcc tgagtgcagg gtctgggtag 17580 cagctgggct tccagctttc tccttgcacc tgacttgggc ttttttctcc tcacaggatg 17640 ggccttgacg gcatgaaaat gcaggtaagg gctgcgggac tgcggctgca tgcttccttt 17700 gcaatcatgt ctccccttta ttatttttcc tttggggttc agaaataact cctcctggac 17760 caggtcccgg cagcgtgcga ctagaggctg agtcagttga ggcctctggc cgtgtccctg 17820 tgggtgctgt tggtctctgt gtgggtgccc accgttctcg atgtctgtct gcagctgtcc 17880 tgtttgcttt ttgccctgat gatctgagtg ggctcagctg tgtaacgaca gacccagagc 17940 tgcagaagct ctcatcttgt tactgtggca ggaggtggct ctggttagtg ggggcttctc 18000 ctccatgcac tcttaattta aggggcttct tcttaaaggt cctgggtgga caggacagga 18060 gcctggagga ccgtggtggc gtgtggccgg gcctgggagc tccccgtgga cttggcctga 18120 gtgggctgga acccagtcat gaggggcacc aagcacaagg agaggggagg ccgggtggat 18180 cctggctgac cctggtcctg tcctggctct gggggccctg tagaccgcag tcctgtccga 18240 ctgggctgag cctgcgcccc tctgtgcgtg tcagaagccc agacagtgtt gccctgtgtc 18300 ttgtggtcta aggagggtta cgccctgcgg tgcctgtctt ctgtccccca cctgattcag 18360 tgtggaaatg tggagtctcc agaggtgtcc tgggtgtcac atttgggatg gatacacgtg 18420 ggcccagcac tgcccgcccc agggctaccc ttggtgccag gtgcccccag ccacgagctt 18480 ttacccagct ggccttgagc tccccagagg ctccccggac actgtccgtg ttttgtgaaa 18540 aggttttcaa aacacatgta aagtggaggt gagtagcaag ccctagagca ggccctggcc 18600 tccctgcccc tccctgtccc ctccctgccc ctccctgccc agcgctccct cagcaccgac 18660 tcatcagtgc acctcaagct gatgagggcg tctgtgtttt gacaaaattg ctctgaggtt 18720 gtcacaccca acaaacttat gacggttcct gagtgtagtc ctcacgttgt ggctggtgtt 18780 tgtgaatcag gattcaggcc aggcctgcac aggccttcag ttgttggtct ttgagctcct 18840 gttagtccag ccgtctctcg tggtctcttt tctcctcctg gaaggtttgt tcctgaaggg 18900 cttcacattg cagatctgac tggttgcttc ttatgttccc tgagtttttg taaactggcc 18960 aggccctgag gctcgatccc attgtgtttc tttggcgaga atgcttttct ggtggtccct 19020 gccttgtccc tccagtgcac gatgtctgga tgcctctgcc acacaccacc ccctgcccag 19080 tccccatgtc tgtctggtca gtgcccagct ctgtctcact agggtttggt caccggccct 19140 ttgaactgag accaggctgt gtacctgtga gcccagctcg gggtgagatt tgaggtggag 19200 ccttcccagc cctgtgcaga attcccatca cctccaggtg tactcagaaa tggggatcat 19260 tggccaggtg cggtggctca cgcctgtaat ccctacactt tgggaggcca aggtgggcgg 19320 atcacaaggt caggagatag agaccatcct ggctaacacg gtgaaacccc gatgctacta 19380 aaaaatacaa aaaaaattag ctggatgtgc tggcaggagc ctgtaatccc agctactccg 19440 gaggctgagg caggagaatg gcgtgaaccc aggaggcgga gcttgcagcg agctgagatc 19500 acgccactgc actccagcct gggcaacaga gcgagacttc atctcaaaaa aaaaagaaat 19560 ggggtcattt ccaggcatca ccatgactga ggtgcgccac tgtcattggg tgagagcagc 19620 tggatgctct atgtgtaggt gctggagcct ctgagggatc gtccagtcct agaagtgtcc 19680 tcagagggac actgtcctgc ctggtggccc atgaagaaag ggagggctcc ctgagtctcc 19740 ctgacgtgtg tctgcctgca gggctcagcc ttctctgagg cccttgtcag ccatgagggg 19800 tgcccagggc tcagagcctg aggctgagcg ttggctgggt gggagccccc acacctggcc 19860 ctcaggcgcc cattggatcc tggaggcagt ggctgggagt gggaggggct gcatctgctg 19920 ctgtaacacc atcctttgtg tgtagggcac caacggctca cagatctggg acaccgcatt 19980 cgccatccag gctctgcttg aggttcgtgg ctccttctct tttctcagcc tcagctgacc 20040 ttcctgtgca cgtaagccca cgcatccacc tgagggcagc actgctggcc acacacttgc 20100 cactcctcga tacttccagt gacctgggct ctggcctctg gcttcagagg gtcgtgctgt 20160 ggagggggcg gccttggcca gcagccttgg gtgttgggct gggtcggggg ccttgggagg 20220 gcaggggctg gaggctgtgt gagaagggga gtctggtgaa ggctgtttct gagagtgcag 20280 gcaggagtgg gactccaggc tcttcttaga actggaactg cttgggccag gcacggtggc 20340 tcacacctgt aatcccagca ctttgggagg ccgaggaggg tggatcacga ggtcaggagt 20400 tcaagaccag cctggccaag atggtgaaac cccgtctcta ctaaaagtac acaaaaatta 20460 gccaagcgtg gtggcgggca cctgtaatcc cagctacttg ggaggctgag gcagagaatt 20520 gcttgaaccc gggaagtgga gggtgcagcg agccgagatt gtgccactgc actccagcct 20580 gggtgacaga gagaggctcc gtctcaaaaa aaaaaaaaaa aaaaaagaac tggaactgtt 20640 tgttatgggc attctcgagc cagtactgga gaaaaacgag agtggatttt tatgccggtg 20700 ggaatgaggt aggtgggatt ctgaaggtgt ttctggagag ccctgagggc tgggccacgc 20760 aaagggcctg cctacacagg gtgctggaga ccctctgggc atggatgctg gccaggcagg 20820 ggggtgctgg catccataaa tggtctcctg cgcccttcca tcttcagtca tatctcatgg 20880 acttttgctg ttttgtcttt aaaggtaagt gcagcaggag accctggcac tctctggaga 20940 tgtctgctgg tttgattctg gtccccggtt ggggcaggat gtggccagga ccatcgggaa 21000 accagcgcag ccatgctggc cgtgcaaggg cagctgagcc tctctgtcct gctgtctctt 21060 ccaggcgggc gggcaccaca ggcccgagtt ttcgtcctgc ctgcagaagg ctcatgagtt 21120 cctgaggctc tcacaggtga ggccggtgcc tggggctctg agggggctga agagggggat 21180 cagggctggg agctcctgca ggcagaagtg cccacctcac ctccaccctg ccctatttcc 21240 tgcactggtg tttcagggtc acccccaccc tcccatcccc tccctagccc ctgctccatc 21300 caccggtcct cctcgggctg gcctcacctg gggcagttct ctgaggcctg cagggtgctg 21360 ggggtgctgg cagtttctgc gtcctgctca tgttggagcc actgtgtgca agggccaggc 21420 acgggcaggg gctgtgtacc ctgagctgca cagcctacac ggcacctcca tgtctctgaa 21480 gcaccttctg cccatggagg tgacgccagc ctgtggactt gccctcctga gactgtttgc 21540 agcaaaagcc ccggtccctc ctgccagatc agctgcccac agaccctgcc cgagcccata 21600 gtttgacctc agtgtctctc acacgtgcct gcaccccagt ctgcagccac agtcatccca 21660 tacatgcgcc ccaacctccc gtgtctccca caccctgtcc cggccacggc ctcagccagt 21720 gtccctctgc ctggaaccgc tgccccccag ccccgtctcc ctcccttcag ctctcactag 21780 gacattgttc tgcagggctt ctgggtcttc ctggcctctg tgtggccaag gctggcaccc 21840 atcttgggct caagcagagg aggggcattg tcctgctgtg cctggcccaa tggcggcctg 21900 ctcctgctcc tgcctcctgc ccaggacttg ctctgggtga tggggacttg gggaggctga 21960 ctgaacccta cggcactcca ggcctcttcc cttctcactg aggtgagaga ggcagccaga 22020 agctgaggtt gttcaggagg cattgggggc gcctggcaca gagcacaccc gcagagacct 22080 gggccccctc cctgccttct ggccggtggg gagatcacag gggagtcagg tgctgactcc 22140 cagtcccgtc tgggctggtt tgagccctcg ctggccagtc acgtttccca gcagctgtgg 22200 gtggtgagct aaacaggtgc aggccctcgc gcgcctcgca gcaccagtgg tggctgtggc 22260 cggcagagta agctcccagg cacgttctgc ctctccagtc ctgcccagtc tgtctcagcg 22320 atgtcccaga tggggacgtc ccgtggtgac gtgttctctg cttccacatt tgccctcgat 22380 gctgcccagg tcccagataa ccctcccgac taccagaagt actaccgcca gatgcgcaag 22440 gtatgcggga gccagcccca tccctgtccc gtcccccagg ggaggccgcc ctcagcaggg 22500 tgggtccttc cctctgaagg gggggctcct ccctggggga ctcctccctt ggcgtttttg 22560 ggtgtcctgc tgtggtggat gcctggccta ggggctcatg cttcatgttg ctgagctgcc 22620 tggcacatgg aggcacagtt ggcttgcaca cacagccgtg cctcagagca gttccagtgg 22680 tcacggcaca cacaggcttc agaaggacag ccgaagtgta gccagtgtgt ccggggaagg 22740 cagaggaaag aagtagacct cagagccggt gtgggctgtg accacaggtg cagactgtga 22800 aattaggcat ggacccagct gctgctgcct gtttacaatg ggggtggggg gcacctgggc 22860 cccatcctgt ccgtcgtgag atctgcaggt gttgagggtg tgagctgcac ccctgagggt 22920 ccctgtgctg gaagctggag gtctgtctgg atgtacccag cttggggccc tggctgcacc 22980 cacacctttg gtggctgggc ccctgccctg accgggtgct ctgtggtggg gagggatgcg 23040 tgcggctgtg gggaggttct gagaactggg gtgtggacac ccccagcctg gagtcatggc 23100 ttgtgctctg cagggtggct tctccttcag tacgctggac tgcggctgga tcgtttctga 23160 ctgcacggct gaggccttga aggctgtgct gctcctgcag gagaagtgtc cccatgtcac 23220 cgagcacatc cccagagaac ggctctgcga tgctgtggct gtggtaaggc tgtggtccca 23280 gcagccccgt ccatacctcg tgtcctgcag atgagctgcg tgctcacttc cactcctgtg 23340 ggctccagcc cagcacacag tccggccagg ccgtaggagc ttgtccttgg atggtgtcta 23400 tatgtggaga actgtgagct ctggctggac ccctaggggc cttgctgggc tgtgtgcaca 23460 gggccctgca ctgcggagct ggtgtccagc ccagccaccg atacttgggg gagccggcgt 23520 ggcccccaag gtttctctct ggtggtttcc actgggtgtc tgaagaggga atttgttggt 23580 gttggttttg gtgccacatc ctttcagcac atctggcttt tgtgtgtgtt tcccagtgga 23640 gaccctgccc ttttctggca gcacagactt ggtttctaag tcatgggcac gtgtgggggc 23700 atgttccctg gtggctgtgc atggaggccc tgacagatga ggttgcagct gctgcttggg 23760 gcacccgagg gcttggttaa cgtggaaatc agctctccgc cccctgttcc tgccccatcg 23820 gttgtcagcc ctagtgttgc ctctagagag ttccgctgtg ccctgggcgc ctgtgtgtgc 23880 tcagcacatg ggcgagttct agggtgctct ctgtgatttc agctgctgaa catgagaaat 23940 ccagatggag ggttcgccac ctatgagacc aagcgtgggg ggcacttgct ggagctgctg 24000 aacccctcgg aggtcttcgg tgagtggtcg gccagcactg cggcgcgcaa acccggggct 24060 ggctagcact gtggtacaca aacctggggg ccagcttttc ccccttgccc gaggctgcaa 24120 gggcccaggt tcaccggcag atctgtctgg agccctccct cagcccaggc tgttctgcgc 24180 tcctccatcc cccggggtgg caggatcctt gtgttgtgga taggagggca tcaggtcaga 24240 cctaggggac agtggagggt tccagtgaga tccacagcct gggctggttc ctgctcagtc 24300 cacagggctt gtgttctgtg gaggctgctg tgtatccaga gcgcctgcag ggaggtgtct 24360 ttggggactg tggggactgt ggggacccat gccatgggca gtaggctgct gtgtgtgcat 24420 ggttgccacc gtactggtct tgggggagga tctcagccct ggtccacctc tgggcacctc 24480 acatacccgc cttcctggtc ccctccacat cacacatggc tttttggggt ggggtcgcag 24540 ctttctgctg tgttcccctc atcttcgctc tcaggtagca caggtgtgtg tcctggacca 24600 gccggcgttt gctctggagg ttggtcaggg aggcagcgtc cgggcccggg ctcactgcaa 24660 cactcttgct tgttgtggct ttgcctgagc tgcagagcct gggcagccag ggtgaaaccc 24720 aacacttggt tcttccctcc ctttcccagg ggacatcatg attgactaca cctatgtgga 24780 gtgcacctca gccgtgatgc aggcgcttaa gtatttccac aagcgtttcc cggagcacag 24840 ggcagcggag atccggtaag gagggtctca gccattcagt gtgggcgctg ccaagtcggg 24900 ggccaagacc cagacgcatc attctgtgac acggccctgg tggcccatct cagaagcgaa 24960 actcatggaa acatgcaaga ggcttcggat gttgtggaat ccagtcatat gccctaaagc 25020 atacaaaata tctgttaggg gctcagaata gcacagttat gatacaaaaa tggattttct 25080 ctctctttta ataatgttaa gaagacatca catacctgac tccaccggtg tcccagaaac 25140 ggtttttaag taacctttcc tgttgaaggg tagcaagtat tcagaaaagt gtacaggttg 25200 gtcttcttga agcaaacagg aagcgaacag tgccagcatt agacatggtg acaccaccag 25260 agccctcggc ccgccccatg acggggccgc ccacatgcct gccaggtcgt gggtgtctgt 25320 tgctcgcttt ggatcttgtc taggtggact cctgaggtgt ggaattcgtg ttgccttctc 25380 ctgctctcct gctctcctgc tcgcggttag tcaggtggct cgggtaacag cagcgttctc 25440 tccctcgggc cttcggttga acacagaatg ccgcgctatc cagctgcctg ttctcagcac 25500 ctgggaggat ttcagtctgg gttattatga agcatctact gtgaacactc ttgtacttat 25560 cttttggggg cacctgggta cccatttctc atggtcacgt acctaggagt ggcattgctg 25620 tgttagaggg tacgttatag ggtatgtgat ttttgtaggt tcttctttat cctatcacga 25680 ttacattttt ttacttttgt tcaacctggt gtagactcac cttggtcaca atgcactgtc 25740 ctttttatat attgctagat tcaatttgaa gaatattttg ttaggatttt agcaactctg 25800 gttacaagag acgctggtct ataatttttt tttctttata atgtttttgt caggttttcc 25860 tgttaagatg atgctggact tagaaaagca gttggaaaat gcttttaaaa tactctttgg 25920 aagaatttat gtaatattca taatatttct gccttaaatg tttgggaaaa attaccggaa 25980 atgccagttg ggcctggaga tttctttgag gaaagttttt aaattagaag ttcaatttct 26040 ttctttcttt ctttctttct tttttttttg agatgggttc tagctctctc actcaggctg 26100 gagtgcggtg taatttcttt aatagtttat aggactgagc agattttcca tttttgtatc 26160 agtctgggga gtcttcccat ttccactcag ctttacactg attcatgcaa agttgttcag 26220 tgtcctctta gatggctctg agcccaacgc tgacatcctc ctcttccttc tgagaatctt 26280 atactgatct tttgaaaaaa aaaaaatctt agtctttgat tctgttttta aagagacttt 26340 atttttggtt tcatcaattt ctattgtttg ttattttctt tctttcttaa tttttttgag 26400 atggagtctt gctctgttgc tgaggttggg gagcagtggc gtgatctcag ttcactgcaa 26460 cctccgtttc cggggttcaa gcgattctcc tgcctcagcc tcccgagtag ctgggactac 26520 aggtgctgac caccatgact ggccaatttt ttggtatttt tattagagac agggttttac 26580 catgttgtcc aagctggtct tgaactcctg acctcaggtg atccaccttc cttggcctcc 26640 cagagtgctg ggattacagg tgtgagccac cacactggcc tttgctattt tctttctcct 26700 ttatttttct aacttgaata cttagatatt tgattttcag gcttttattg aaatatgaat 26760 ttgaggctat aaatgagttt tgagatatca ttcagttaaa tgtgtgttct ggtgcttgct 26820 gtggtagcac agatactaaa agtgttttct gtttctactg ttcttctctg gcccatgagt 26880 tatgtgggag tatgctgctt catttacaat ctgagaatgt tctggtgtgg tttttttgga 26940 agccgtggat ggagcagggg ttttcttgtg cttcacaggt gcagctagga gggcactgtg 27000 tccagggtct tctgtcggcc tggcgtggcc cttggccatg tgctgctctg cggcatgagg 27060 tgggcgtgag ttgtcctcag ccacatttag agaattggcc ttttaaaaaa tagatcatct 27120 tttaaaaatc actgtaataa aagtaaagca ggttctttgc aaacaagact tgcaaaatac 27180 agagaagcgc aaagaagaag ctaagtcgcc cctcctcgcc cctgaaggag aatctgctgt 27240 tgctgtttgg tctccacatt tccatggcgg cttgctgccc ctttcacgcc tggcccactt 27300 tgtgcctggt gaggtttcta aaagccccac ccttgagcgc gctcctccag cacgagcagt 27360 aatggcacag gtgttgtgtc attttactca gtagcctctg ggttattttt cagttttcct 27420 tgttgttttt tagcttttcc ccattttaac cttaactggt attttcttgt taaatattta 27480 ttcatgacca ttattattcc ctagagccac atggcttggg gtccacctgc ctgggtccgc 27540 ccccatccct gccccttctg gctgtctgac ctggcctggt gacttctctt ctctgctcat 27600 ctctctccct gcctgagtgg gcaagagtac agcctcacag agtggtggga ttgtgtgaga 27660 tgccacaggg aagcacatgt cagttgttgt cactgtgtag aacaatgagt cccggatgtg 27720 gcccgcaggg gagcaatggt gacttaatcg cgggcttcct ctgcatttct ttggtgactt 27780 ccaagctaga acattctttt tttgtttatt tgtttgaagc agggtctcac tctgttacct 27840 aggctggagt gcagtagcaa aatcatggct caccacagtc tcaaacttcc gggctcaagc 27900 aatcctccca cctcagcctc ctgagtagct gggactacag gtgcatacca tcacctgtgg 27960 ctaatttttt aaatgttttg tattttttaa atgttgctca ggctggtctt gaactgctgg 28020 gctcaagcaa tcctcccacc tcggcctccc caaatgctgg gattacagag tgagccacca 28080 cacccagcca tttttaaaat tttcaccagg aagttttttc tttcattttt aagcacagta 28140 agtatttgtg tattatgtta cagatatttt cccctcaatt tctttgttct ttttatctct 28200 ttagggagta tgaacataag tttttaactt ttaaatggtt aaatatatta gtgtgatttt 28260 tatattaaga ttttatttta tttatttttt ttttttttga gacggagtct cgctctgtcg 28320 cccaggctgg agtgcagtgg cgggatctcg gctcactgca agctccgcct cccgggttca 28380 cgccattctc ctgcctcagc ctcccaagta gctgggacta caggcgcccg ccactacgcc 28440 cggctaattt tttgtatttt tagtagagac ggggtttcac cgttttagcc aggatggtct 28500 cgatctcctg acctcgtgat ccgcccgcct cggcctccca aagtgctggg attacaggcg 28560 tgagccaccg cgcccggcct atattaagat tttaaacttg ccgggcgcag tggctgacgc 28620 ctgtaatccc agcactttgg gaggccgagg cgggtggatc acaaggtcag gagatcgaga 28680 ccatcctggc taacacggtg aaaccttgtc tactaaaaat acaaaaatta gccgggcgtg 28740 gtggcgggcg cttgtagtcc cagctactcg ggaggctgag gcaggagaat ggcgtgaacc 28800 cgggaggtgg agcttgcagt gagccgagat ggtgccactg cactccagcc taggcgagag 28860 tgcaagacac cgtctcaaaa aaaaaaaaaa gattttaaac ttacctggag agtttttgag 28920 atacagtttg gagttgcaag ttactttaac actatttata tggaatattc tattttacta 28980 gacagactta aattctccct taaattcaca aatttataga aaagttacaa aaatactgaa 29040 aagtgctcct gtttactctg actagaattc tttagtgggt ggcaccctac cctgagggct 29100 tcatgacctg tcctcccaca tgatccaggc tctaccctca gggcttcatg acctgtcctc 29160 ccacatgatc caggctctac cctcagggct tcatgacctg tcctcccacg tgatccaggc 29220 tctaccctca gggcttcatg acctgtcctc ccacatgatc caggctctac cctcagggct 29280 tcatgacctg tcctcccacg tgatccaggc tctaccctca gggcttcatg acctgtcctc 29340 ccacgtgatc caggctctac cctcagggct tcatgacctg tcctcccacg tgatccaggc 29400 tctaccctca gggcttcatg acccttcctc ccacatgatc caggctctac cctgaggact 29460 tcatgacctg tcctcccacg tgatccaggc tctaccctca gggcttcatg acctgtcctc 29520 ccacatgatc caggctctac cctcagggct tcattacctg tcttcccaca tgatccaggc 29580 ccattctttc ttgaaccatt ggaaaggaat ttgcagatag gatgtgtacc cctaactgcc 29640 tgagtatttc ttagcaggtg tattcttttg tgcaagtgta agtcagaatg ttaatgttga 29700 tgaaatacta atctgcaggc ctaattgttc caataatgtc ctttatggca aaatcttccc 29760 ctcaagcttt aatccaatat catgggttgc atttgtttat ttttaattat ttttcttttc 29820 tttttctgtt ttccttaccc ttctcactgt gcacatgggt tgcatttagt tatcacattg 29880 acaccctttt aacctggaat aattccttaa tctttccttg tgtttgatga ctttgtcatt 29940 tttgaattgt tccacaagtt attttgtaga atatcctcag tgtttttttt tttctggtgt 30000 ctcgtgatta gattcaggtt atgaaactac atttttgtca ggaagattgc agaagaaatg 30060 gggccttctc ctgcacctta ccaggaagca cacaccaact ttgatccctt gattaaggtg 30120 atgaccgctg tacttagttt ctccgctatg aagttgcttt ttttttcttt gtggggagac 30180 agttttagac tatgtaaacg tcctatttct catcatactt atacctgtta gtgttagcat 30240 ttgatgatga ttcttgcctg aatcaattat ttgcataatg cttacaaaat tatcattcct 30300 tccatatgta ttagttggtg ttcttccaaa agcttttcct ttgcctctgt ttatttactt 30360 atttatcact gtggacgcat agattcttac gcaattgatt ttgatactga tctcatagct 30420 agacaatttt gctaaacttt taaaaaaatt tatgtacttt atcttttata gcagctttaa 30480 atttacagaa aatttgagtg gaagatgcag tgttcccata aagccgctaa ctcctcgcac 30540 cttccctcaa gtttccccag tactaacatc ttgcattcaa gtggtgcgtt tgcaacattc 30600 ataaattatt atcgtccaga gtccattgtt tacattcagc ttcctcttca tgttgttcat 30660 tctgtggttt cacagatgtg tgatgcatgt gcccaccact gcagtgtcac acaggatctc 30720 actgccccgg agtcctctgc gctgtccccg cctccagaac cccttagtag caaacactga 30780 tatttttact gtctccatag ttttgccttt tcagactgac ctatttcact tagtaagaag 30840 catttaagat tcctgagtct ctttctatgg ctcaatagca catttctttt tagtgctgaa 30900 taatattcca ttgtctggat gtaccacagt ttattcattc acctactaag gtgaatgtct 30960 tgcttgcttc caagttttgg caactatgaa taaagttgct atcaatgtta gcgtgcacat 31020 aagttttcag ctcatttggg taaatgccaa gaagcatgat tgcgggatcc tatggtaaga 31080 gtgtgtttag ttctgtaaga agctgccaaa ctgtatctta agtggctgca ccatttgcgt 31140 ttccaccagc aatgatgagc gttttgttgc tccacatcct caccagcatt tgctgttgtg 31200 ttttgggttt tagcctttct aagaggtgtg tagtggtatc tccttgtttc aatttgcaat 31260 tccctaatga cattatgtta aaatcttgtc atatagttat ttgccatctg tgtatctttt 31320 tcagtgatgt gtcctttaaa gtctttggct catttttaaa ttaaattttc ttattgttga 31380 gttttagttc ttcatatatt ttggctgcca gtcctttatc agatatgtct ttcgcaaata 31440 ttttctgcct gtgtcttgtc ttttcattct attaacagta tcttttgcag agccagtttt 31500 catttcaagg aagtccagct tatcaatgtt ctctttcatg tatcatgttt ttggtgttgt 31560 atctaaaaag ttactgccaa gcccaagggt acctagattt tttcctgtgt tatattctag 31620 gatttttaaa gttttgcatt ttacatctag gtccatgatt cattttgagt taacttttgt 31680 gaagggttta tggtttgtgt ctagattttt tttttttttt tttttttgca tgtggatgtc 31740 cagttgtttt ggtaccatct gtcaagaaga ctctttttgg gtcattttgt tgcctttgtt 31800 tctttgtaaa aaatcagttg actgcatttg catgggtcta tttctgagct ctctgttcca 31860 ttgcattgat ctgtttgttc ttctcagcaa tcccacactg tcttggttcc tgtagctctg 31920 tagtaggcct tgcagtcagt taccgcccct gttctcactt cagtgttctc ttcaatagtg 31980 ttttgactat tctaggtttt ttccctctcc atatacattt tagagtcagt ttgtcaatag 32040 tttacaaaat aacttgctga gactttgatt gggattacat tgaatctgta gctcaagttg 32100 gaaagatctt ttatttcttt catcagaatt ttgtagtttt catcatatat agatcttgta 32160 catattttgt tgtttatacc taaggatttc atttttttgg tgctaatgta aatggcgttg 32220 tgttttaaat gtcaaaatct aattgttcat tgctggtagg aaaacaactg accctttttt 32280 ttttttttaa gggacgcagt cttactctgt tgcccaggca gagtgcagtg gtgccatcat 32340 agctcactgc agcctcaaac tcctgggctt aaggaatcct cctgtctcag cctcctgagc 32400 agctaggacc acaggcatgt gccactacgt tcagctaatt tttcaatttt tttgtagaga 32460 tgggatcttg ctctgttgcc caggctggtc tcaaactccc gtctgctttg agatgattat 32520 atatttgtgt cctttgttaa tttagaggat tattatggat ttttctaatg ttaagacacc 32580 tttgtatttc tgagatcgac cttagtattg gtctatattt aagacagtat tcagtttctc 32640 agttgttttt tgttttttgg tttttttttt tgagacagag tctctgtctc ccaggctgga 32700 gtccagtggc acaatctcag ctcaccgcaa gctctgcctc ccggattcac gccattctcc 32760 tgcctcagcc tcccgagtag ctgggactac aggcgcctgt catcatgccc agctaatttt 32820 ttgtattttt agtagagacg gggtttcacc atgttagcca gggtggtctc aatctcctga 32880 cctcgtgatc tgcccacctc gatctcccaa agtgctggga ttacaaggcg tgagccactg 32940 cgcccggcag cagtttctca gttttaattt ggagttttgc atctgtgttc atgagtgagc 33000 ctgaaatttt cacttttcca tatcttattt ctctgggttc ctagaatgag ctagagagtg 33060 ttcctccttt ctgttctctg gaagagtttg tgtgagatta gaatgagtgt gtctgataat 33120 ttagttgcat tcatttataa aattcctagg cctagagttt tttttctggg aaaagtttac 33180 attttgactc atttttttag tagttttagg actgtttagg ttctctattt cttgattgag 33240 ccagttttga taagttaatc tttctaattt gtagatattt tctctaagtt tgcaaatgta 33300 atacataaaa ctttcttgtc atttctcacc atatctgtag ttctatcttt ttattgctaa 33360 tattactaat ttgtactttg actatttgta tttgttacct gttgccgagt aacaatatta 33420 gtacaaacct agtggcttag aacaacacac attgattact tcaccgtttc tgtgtgtcag 33480 aagtccaggc gcggcctcgc aggtcgtcct ctgcctcagg gtctctccgg gcttcagtca 33540 gggtgttagc caggaccggg gtctcgcctg agcttccagt gaggaaggat ctgcctctga 33600 gcacacaggg tcctcggcac gatcccattc ctcagctgga agctgccgac tgccgtctgc 33660 tgcggggcct ctctagatgg catcttcaca aaagcgagaa gggagagttg gtagagggag 33720 tctgctagca ccatgggagt cgcggtcaca cagacctcgg tcccaggacc cgcacccatc 33780 aaccctgccg tgatctgctg gttaaagaca agtcccacgt cccacagggt gacactggag 33840 tagacacttc gctctggcct tttcagagaa ctggttattt tttggaaata tcagttagat 33900 gtaggatggg tcttgtcttc taaatctatt gttttctctc taattgattt tttcctgttt 33960 ttatttagtt cactttgttg ggtttgctca agcctgggtc actggatctc agggatgctg 34020 ctcctgtttg cagctgtgtc tgcaggggct tcccaaggcc ttgctttccc ctcacgtccc 34080 tttctcagac tctgccaatc cgcttcccgc tctggtgtcc tgtggttgct tctttttaaa 34140 accctcatcg gtctgtgtaa actgtttatt tttatgtggt ttttaaggga gaccattctc 34200 attcttttga gaccctggaa aggatggaat tgggataggt aaactgctgt tttaccagaa 34260 tgttcactgg accaatctcg tgttccaggg agaccctcac gcagggctta gagttctgtc 34320 ggcggcagca gagggccgat ggctcctggg aagggtgagt gagcctccac tcgtgagtgc 34380 agagatgcat gggatccaga ggtttctgct ctcacacact gcgttcataa atgttggctt 34440 gtatgttgtt gctacaccag aagtttctgg aagtgagctg ccagcccgtg acttctgggg 34500 gacctcgttc ctttgtggca tgcgtggcct ttgccccggt ggaaattgct cagtacgttg 34560 ctgggcgcag ccgggctgct gggagcgcgc tgtagcctga gcgtggctat tccctccacc 34620 ctttctgctt gctcttaggg tccagcagac agagctgctg tcttccacgg ccttaatgcc 34680 tgaggcactg gagttggtgg gctggctggg gcacgtgtga ttgttgcaga atgcgtgttg 34740 tttcacacac cggctgtgaa cagggtggaa gggctgaggc tctccctgtt tccctccagc 34800 tcctggggag tttgcttcac ctacggcacc tggtttggcc tggaggcctt cgcctgtatg 34860 gggcagacct accgagatgg gtgagtgagt gcctgtcctc tggtgggtgg gggttctcaa 34920 cccaatgctc tgtcatgagt gttttttgct ttgacatttg gttttagggt ttgtttgttt 34980 gtttgtttgt ttttgagacg gagtctcgct ctgtcaaccg ggctgacatg cagtggcatg 35040 atcctagctc actgcagtct caaactcgtg ggctcaagcg atcctcccga gtagctggga 35100 tcacaggtgc acgccaccac cccgggctaa tcttttaaaa cttttatgta gagatggagt 35160 cttgctgtgt tgctcacact ggtttgggct caagcagtct tcctacctcg gccttccaaa 35220 gtgctggggt tacaggcatg agccaatgtg cctggcctgt ttttaatatt tttaaacagt 35280 gagataagat ccccggttga aatgaagatg tttccctggt cccacagctc tctggagctt 35340 cctgacatgt atgctggagg gacgcttctg gtctccggcc cctccaggca tacagatgcc 35400 tcccaaccct gagtaggaag attagggtcc acggcctcgc tggagcgggt tagaaggcag 35460 gagatctccg gtcccagccg tgtctccagc cgccggactc tctcccagcc ctgtctccag 35520 ctgccccact gtctcccaga gtctgccgtg tggatgttta gaggtgggga gcaccgtgct 35580 tggctgagtg cagcttgtga gacgctgctc ccaagcactg cagacctcac tcagcctgac 35640 gcgtccgtga ggccatcctc ggtactcgca tgtccctttg tcttcccagc gactctggga 35700 ggcaggagta tctgttccca gttcacatct gcaaaagtca agctcgggtt tcagtagtgg 35760 cccatggccc ttaggtaggg tggccccatc gtgcaggctc ctccccgtac cccaaggcag 35820 cctgctgggg tgagaagcca ggggtctggg accttccttg gtgtgatggt gtctcctgtc 35880 tctggtcttt gcaggactgc ctgtgcagag gtctcccggg cctgtgactt cctgctgtcc 35940 cggcagatgg cagacggagg ctggggggag gactttgagt cctgcgagga gcggcgttat 36000 ttgcagagtg cccagtccca gatccataac acatgctggg ccatgatggg gctgatggcc 36060 gttcggtggg gacgacggga ccgtccctga gccttgggtt tgggtagagg agggacactc 36120 agctgtgagc cggtggcctg ggctgagtga atgtagagag gaggggaggc ctgtgggcca 36180 ggtcagctgc cactctggga acagacacct acaagagcca catgcctggt tcctggggca 36240 agaacgtggg ctgctctgac caagtggggc cctgcagaga ggctcgcctc ttagaagtga 36300 accacccacc attagccatg tcagtggaag agcaagcaca tcagggaccc atggaaacag 36360 cgaggtgggc tgcgatgagg atgctgcttc ctggtgtggt agtgatgacg gtcacagcag 36420 ctgctctctg tggccctact gtgttcacag ctggtgctga gccacatatg tgccaggtgc 36480 acacacacgc agacgcatgc aggcaggcat cagtgtacac actgatgtgc acacacagat 36540 gtacatggag acagatgcac acacaggcct atgcacacac gtacgcatgc ccacacaggc 36600 acctgtgtcc acacacatac agatgcaccc acagcatccc atctgtgcca cacactgaca 36660 taggtacatg gagacagatg cacacacagg tctgtgcaca cacgtatgca tgcacaggca 36720 cctgtgtaca cacacgtaca gatgcaccca caggatccca tctgtgccac acacagacgt 36780 aggtacatgg agacagatgc acacacaggt ctgtgcacac acatacatac gcatgcacag 36840 gcacctgtgt acacacatgc agatacaccc acagcatccc atctgtgcca cacacagaca 36900 taggtacatg gagacagatg cacacacagg tctatgcaca cacatacgca tgcacaggca 36960 cctgtgtaca cacacgtaca gatgcaccca caggatccca tctgtgccac acacagacgt 37020 aggtacatgg agacagatgc acacacaggt ctgtgcacac acatacatac gcatgcacag 37080 gcacctgtgt acacacacgc agatacaccc acagcatacc atctgtgaca cacacagacg 37140 taggtacatg gagacagatg cacacacatg tctgtgcaca cacatacata cgcatgcaca 37200 ggcacgtgtg tacacacatg cagatacacc cacagcatgc catctgtgac acacacagac 37260 gtaggtacat ggagacacat gcacacacag gtctgtgcac acacatacgc atgcacaggc 37320 acctatgtac acacatgcag atacacccac agcatcccat ctgtgccaca cacagacata 37380 ggtacatgaa gacagatgca cacacaggtc tatgcacaca cgtatgcatg cacaggcacc 37440 tgtgtacaca catgcagatg cacccacagt atcccatctg tgccacacac agacatacgt 37500 acatggagac agatgcacat acaggtctat gcacacatgt acacatgcac aggcacctgt 37560 gtacacacat gcagatgcac ccgcagtatc ccatctgtgc catacacaga catacgtaca 37620 tggagacaga tgcacataca ggtctatgca cacatgtaca catgcacagg cacctgtgca 37680 cacatatgca gatgcacccg cagtatccca tctgtgccac acacagacat acgtacatgg 37740 agacagatgt acacacaggt ctatgcacac atgtacacat gcacaggcac ctgtgtacac 37800 acatgcagat gcacccgcag tatcccatct gtgccacaca cagacatacg tacatggaga 37860 cagatgcaca cacaggtcta tgcacacatg tacacatgca caggcacctg tgcacacata 37920 tgcagatgca cccgcagtat cgcatctgtg ccacacagac atacgtacat ggagacagat 37980 gtacatacag gtctatgcac acatgtacac atgcacaggc acctgtgcac acatacatac 38040 agatgcaccc gcaacatccc gtctgtgctg ccctattagg tttgtggcca tttggggaat 38100 cttcctaaaa ccctaaaagc tagggcaggt ctgcttgagc aggagcagca gggtctgggg 38160 gacccctgag ggcaggacag tcagggaccc acagttgagc tgggcccgct gagccctgga 38220 tccttcttgg tgtcttatcc tggccagcaa gcaagtgtga gctcctgtgg gtctccagag 38280 gcccatgagg accagtgggc cagttgggaa caaggcttgg cgtcctcttc aggggggaac 38340 accagggcag gcctgaggag gcctgtgtcc ccagcctgtc attgctgtgg ctccgcttct 38400 cagggagcct aggaagaagg tgtggcaaga gcccgaggcg ctggctgcac ctggcggggc 38460 ctgtgggcgt cagtttagac ccatccattc tcactgcagc attccagggt ttgcccttat 38520 gctcggctgt gtgagggtga ggatgatgct gtgggggcat gcatgctggg tgtgtttcag 38580 ccttctcttc caccaggcat cctgacatcg aggcccagga gagaggagtc cggtgtctac 38640 ttgagaaaca gctccccaat ggcgactggc cgcaggtatg ccgccaggga cctgagcgca 38700 caaggcccag cactgacctc cagcgtgcat ggctgtttcc acgtccccct gctctgtgtc 38760 ctttttgggg tactttggac acttgggagg cgtcacctct gccagtgaat gccacagttg 38820 gtggcaggtc tgtggcaggt ggtcgggtcc taaagtccag atcttgctgt tgtttcaagt 38880 gatgctctgg gtgggggagg agctggatgg gagaagccag tgggcgggaa gcctttttgc 38940 tgcaggacag accctcccac tccagatgac ctagtggccc ctcactgagc cagaagtccc 39000 tgtggtgtgg gtgtcatgag gtcatgtgag gccaaccgcc ctcccctggg atgaggctga 39060 gttggtggaa gctgatgtgg ttgtgagggg ctggtgaccc tggcttaggg tttgctgcag 39120 ggcggggagt ctgagctggg ctgatggtgc catgactgat gcgggatgga ctacttgctt 39180 tcctatgctc ttgcttaatt agccctttcc aggctgactc acccacaagc cagccaagcc 39240 aacagccagg gctccagttc agggactagc cctcagctga ctggtgaagc ctttgtgttt 39300 atttctctgt gttcttttag gaaaacattg ctggggtctt caacaagtcc tgtgccatct 39360 cctacacgag ctacaggaac atcttcccca tctgggccct cggccgcttc tcccagctgt 39420 accctgagag agcccttgct ggccacccct gagaacatgc ctacctgctg ggtgccgtct 39480 gtgcgttcca gtgaggccaa ggggtcctgg ccgggttggg gagccctccc ataaccctgt 39540 cttgggctcc aacccctcaa cctctatctc atagatgtga atctgggggc caggctggag 39600 gcagggatgg ggacagggtg ggtggcttag actcttgatt tttactgtag gttcatttct 39660 gaaagtagct tgtcgggctt gggtgaggaa gggggcacag gagccgtgac ccctgaggag 39720 gcacagcgcc ttctgccacc tctgggcacg gcctcaaggt agtgaggcta ggaggttttt 39780 tctgaccaat agctgagttc ttgggagagg agcagctgtg cctgtgtgat tccttagtgt 39840 cgagtgggct ctgggctggg gtcggccctg ggcaggcttc tcctgcacct tttgtctgct 39900 gggctgaggg acacgagggc aaccctgtga caatggcagg tagtgtgcat ccgtgaatag 39960 cccagtgcgg gggttgctca tggagcatcc tgaggccgtg cagcagggag ccccatgccc 40020 ctgggtcgtg agcttgcctg cgtatggggt ggtgtcatgg agcctcatgc ccctgggtcg 40080 tgagctcgcc tgagtatggg gtggtgtcat ggagccgcat acccctgggt tgtgagctcg 40140 cctgcatatg cagggtctgt catggaacat cccaagtctg tgcagcaggg gagccccatg 40200 cccctgggac atgaacccac ctgcgtggaa tgctgtttgt gaggtgtcta cagggtttat 40260 agtagtcttg tggacacaga aatgcacagg ggacacttac ggacacagaa atgcacaggg 40320 gaggccgagc ataaccaggg gtgaggggca ggcagcagtt gtagttactg ccgcggggca 40380 ctgctatgtg cagggacagc cagcgcccag cccatcacca ctccctgggc tggctggcag 40440 gtatggcacc ctgggagccc ggcatatacc cagggcaccc ctacggctgc cgccagtctc 40500 atgcccaggt gggtgctctg ggctggagcg agggccaggt tttgggccga ggcttcccca 40560 ggcaatcctg tgagctccct tctagcctct gacccagtct ggtctggctt gcatggatgt 40620 agggcttggg gtgggaagtt caggtcctgg ctttgccttt gcctgatgtg gatgagcagc 40680 tcacatgctc agggccacct gagactgtca ctgctctccc ctggctactg ggaggagtca 40740 ctgagagctt cgttacccct gctgccttgc ccagggcaca ccctatacct cctcatctgc 40800 tcttcccctc cctgccgcct tctgggcagg tagcagtccc tggcctctcc ccctggctga 40860 tcactctccc tcaggcagtg gagatctgcg tctggacacc ctcagatcct gtcattgcct 40920 gcccagagtc cttcaggggc acccctctgc cttggtgtgc ggtccagggc tctcacccag 40980 gtgccgcacc ctctggggtc ttctgtccag ctcccttgcc ccatgtgctg tcactgactc 41040 tccttgggac tcgcctgcct gctcagagcc ctgcagggct tggtcagctg cctgttcagt 41100 gtcaacactt ccctgcacat cttaaaactg ggctttattt tcgctgaagg aactgtgttg 41160 ggacccttga catctgtcag gtttgcacat gctgtttttt tttctcagcc cacgtgttct 41220 cccccacgtg gggtagcagc aggacagaca gtgaatcaca gagtctgccc tgagcagagg 41280 ctgctgtccc tgggactcct agccatggtc agactgtaca aaacggtttt ccagaaatga 41340 aatgtaaatc catttttata ctgaaaatgt tactgaaagt cacttttatg agcatctgcc 41400 ttaataaaca gacattgatt cccttatcag aagcctgtca cactgtgttt cgtttcatcc 41460 tggggagaac tgcagatttg gggtttctgg ctgtcatacg tcacctgcct gtggggcgag 41520 tgggaggccc agcctggttt agggaacaag agtgacgtga ggagtagcag ggtgcgtctc 41580 cagttacctg agggaaaaca gatattttaa gagataatag catagcctat tttaatatgt 41640 tttaaaggcc ataagcatat ccaggaagat aaataaacgt gatacaatgt ccacatagga 41700 ggaactttct ttcactgcat tgttttcctt cacagtggcc ttcaagtcac aggacgcagc 41760 gattccctgc cctcttcggt gttattacac aggcaggact tcagtgtcag tatccctgcc 41820 ttcagtcttc tttagaaatc acatctgtgt tcaatccatt gtttagaggg agtgtatttt 41880 tcctgttcca cgaagaggac tttttgttca caattggatc acaatgcaga ggagtctgtt 41940 cctcccccgt cggcttctcg gtgctgggag ggtgacctgt cccagatgac tcatcaccct 42000 gacatgctct tgacaaagga caccaccaag aggagatggc agctgtaccg gtgcagcctc 42060 tgtctgaggg ggatatttgc ctcagtgtga ttaaaaatca gtcatgaaag atttttgaat 42120 tcagattatt tttatcagga acagattttg aacatcctga aatcttttcc ctggcatcat 42180 attaggtttt ctttgttcac tatgatgtaa agtttcagac tcttgatatt tttaatatca 42240 acatagacgg taggacaagg aacggtacca gaaatgagta aagagacaat aatgataaga 42300 tcgatttatc aagacataac aaccccaaat gtatatgcac taaataacag cttcaaaata 42360 catgaagcaa aatggcagaa ttgaagagaa tgagataaaa acagaatttt aacgggtgct 42420 ttccgtactt tgtaactgac agacatgaga 42450 4 732 PRT Homo sapiens 4 Met Thr Glu Gly Thr Cys Leu Arg Arg Arg Gly Gly Pro Tyr Lys Thr 1 5 10 15 Glu Pro Ala Thr Asp Leu Gly Arg Trp Arg Leu Asn Cys Glu Arg Gly 20 25 30 Arg Gln Thr Trp Thr Tyr Leu Gln Asp Glu Arg Ala Gly Arg Glu Gln 35 40 45 Thr Gly Leu Glu Ala Tyr Ala Leu Gly Leu Asp Thr Lys Asn Tyr Phe 50 55 60 Lys Asp Leu Pro Lys Ala His Thr Ala Phe Glu Gly Ala Leu Asn Gly 65 70 75 80 Met Thr Phe Tyr Val Gly Leu Gln Ala Glu Asp Gly His Trp Thr Gly 85 90 95 Asp Tyr Gly Gly Pro Leu Phe Leu Leu Pro Gly Leu Leu Ile Thr Cys 100 105 110 His Val Ala Arg Ile Pro Leu Pro Ala Gly Tyr Arg Glu Glu Ile Val 115 120 125 Arg Tyr Leu Arg Ser Val Gln Leu Pro Asp Gly Gly Trp Gly Leu His 130 135 140 Ile Glu Asp Lys Ser Thr Val Phe Gly Thr Ala Leu Asn Tyr Val Ser 145 150 155 160 Leu Arg Ile Leu Gly Val Gly Pro Asp Asp Pro Asp Leu Val Arg Ala 165 170 175 Arg Asn Ile Leu His Lys Lys Gly Gly Ala Val Ala Ile Pro Ser Trp 180 185 190 Gly Lys Phe Trp Leu Ala Val Leu Asn Val Tyr Ser Trp Glu Gly Leu 195 200 205 Asn Thr Leu Phe Pro Glu Met Trp Leu Phe Pro Asp Trp Ala Pro Ala 210 215 220 His Pro Ser Thr Leu Trp Cys His Cys Arg Gln Val Tyr Leu Pro Met 225 230 235 240 Ser Tyr Cys Tyr Ala Val Arg Leu Ser Ala Ala Glu Asp Pro Leu Val 245 250 255 Gln Ser Leu Arg Gln Glu Leu Tyr Val Glu Asp Phe Ala Ser Ile Asp 260 265 270 Trp Leu Ala Gln Arg Asn Asn Val Ala Pro Asp Glu Leu Tyr Thr Pro 275 280 285 His Ser Trp Leu Leu Arg Val Val Tyr Ala Leu Leu Asn Leu Tyr Glu 290 295 300 His His His Ser Ala His Leu Arg Gln Arg Ala Val Gln Lys Leu Tyr 305 310 315 320 Glu His Ile Val Ala Asp Asp Arg Phe Thr Lys Ser Ile Ser Ile Gly 325 330 335 Pro Ile Ser Lys Thr Ile Asn Met Leu Val Arg Trp Tyr Val Asp Gly 340 345 350 Pro Ala Ser Thr Ala Phe Gln Glu His Val Ser Arg Ile Pro Asp Tyr 355 360 365 Leu Trp Met Gly Leu Asp Gly Met Lys Met Gln Gly Thr Asn Gly Ser 370 375 380 Gln Ile Trp Asp Thr Ala Phe Ala Ile Gln Ala Leu Leu Glu Ala Gly 385 390 395 400 Gly His His Arg Pro Glu Phe Ser Ser Cys Leu Gln Lys Ala His Glu 405 410 415 Phe Leu Arg Leu Ser Gln Val Pro Asp Asn Pro Pro Asp Tyr Gln Lys 420 425 430 Tyr Tyr Arg Gln Met Arg Lys Gly Gly Phe Ser Phe Ser Thr Leu Asp 435 440 445 Cys Gly Trp Ile Val Ser Asp Cys Thr Ala Glu Ala Leu Lys Ala Val 450 455 460 Leu Leu Leu Gln Glu Lys Cys Pro His Val Thr Glu His Ile Pro Arg 465 470 475 480 Glu Arg Leu Cys Asp Ala Val Ala Val Leu Leu Asn Met Arg Asn Pro 485 490 495 Asp Gly Gly Phe Ala Thr Tyr Glu Thr Lys Arg Gly Gly His Leu Leu 500 505 510 Glu Leu Leu Asn Pro Ser Glu Val Phe Gly Asp Ile Met Ile Asp Tyr 515 520 525 Thr Tyr Val Glu Cys Thr Ser Ala Val Met Gln Ala Leu Lys Tyr Phe 530 535 540 His Lys Arg Phe Pro Glu His Arg Ala Ala Glu Ile Arg Glu Thr Leu 545 550 555 560 Thr Gln Gly Leu Glu Phe Cys Arg Arg Gln Gln Arg Ala Asp Gly Ser 565 570 575 Trp Glu Gly Ser Trp Gly Val Cys Phe Thr Tyr Gly Thr Trp Phe Gly 580 585 590 Leu Glu Ala Phe Ala Cys Met Gly Gln Thr Tyr Arg Asp Gly Thr Ala 595 600 605 Cys Ala Glu Val Ser Arg Ala Cys Asp Phe Leu Leu Ser Arg Gln Met 610 615 620 Ala Asp Gly Gly Trp Gly Glu Asp Phe Glu Ser Cys Glu Glu Arg Arg 625 630 635 640 Tyr Leu Gln Ser Ala Gln Ser Gln Ile His Asn Thr Cys Trp Ala Met 645 650 655 Met Gly Leu Met Ala Val Arg His Pro Asp Ile Glu Ala Gln Glu Arg 660 665 670 Gly Val Arg Cys Leu Leu Glu Lys Gln Leu Pro Asn Gly Asp Trp Pro 675 680 685 Gln Glu Asn Ile Ala Gly Val Phe Asn Lys Ser Cys Ala Ile Ser Tyr 690 695 700 Thr Ser Tyr Arg Asn Ile Phe Pro Ile Trp Ala Leu Gly Arg Phe Ser 705 710 715 720 Gln Leu Tyr Pro Glu Arg Ala Leu Ala Gly His Pro 725 730 

That which is claimed is:
 1. An isolated peptide consisting of an amino acid sequence selected from the group consisting of: (a) an amino acid sequence shown in SEQ ID NO:2; (b) an amino acid sequence of an allelic variant of an amino acid sequence shown in SEQ ID NO:2, wherein said allelic variant is encoded by a nucleic acid molecule that hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (c) an amino acid sequence of an ortholog of an amino acid sequence shown in SEQ ID NO:2, wherein said ortholog is encoded by a nucleic acid molecule that hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; and (d) a fragment of an amino acid sequence shown in SEQ ID NO:2, wherein said fragment comprises at least 10 contiguous amino acids.
 2. An isolated peptide comprising an amino acid sequence selected from the group consisting of: (a) an amino acid sequence shown in SEQ ID NO:2; (b) an amino acid sequence of an allelic variant of an amino acid sequence shown in SEQ ID NO:2, wherein said allelic variant is encoded by a nucleic acid molecule that hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (c) an amino acid sequence of an ortholog of an amino acid sequence shown in SEQ ID NO:2, wherein said ortholog is encoded by a nucleic acid molecule that hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; and (d) a fragment of an amino acid sequence shown in SEQ ID NO:2, wherein said fragment comprises at least 10 contiguous amino acids.
 3. An isolated antibody that selectively binds to a peptide of claim
 2. 4. An isolated nucleic acid molecule consisting of a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence that encodes an amino acid sequence shown in SEQ ID NO:2; (b) a nucleotide sequence that encodes of an allelic variant of an amino acid sequence shown in SEQ ID NO:2, wherein said nucleotide sequence hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (c) a nucleotide sequence that encodes an ortholog of an amino acid sequence shown in SEQ ID NO:2, wherein said nucleotide sequence hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (d) a nucleotide sequence that encodes a fragment of an amino acid sequence shown in SEQ ID NO:2, wherein said fragment comprises at least 10 contiguous amino acids; and (e) a nucleotide sequence that is the complement of a nucleotide sequence of (a)-(d).
 5. An isolated nucleic acid molecule comprising a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence that encodes an amino acid sequence shown in SEQ ID NO:2; (b) a nucleotide sequence that encodes of an allelic variant of an amino acid sequence shown in SEQ ID NO:2, wherein said nucleotide sequence hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (c) a nucleotide sequence that encodes an ortholog of an amino acid sequence shown in SEQ ID NO:2, wherein said nucleotide sequence hybridizes under stringent conditions to the opposite strand of a nucleic acid molecule shown in SEQ ID NOS:1 or 3; (d) a nucleotide sequence that encodes a fragment of an amino acid sequence shown in SEQ ID NO:2, wherein said fragment comprises at least 10 contiguous amino acids; and (e) a nucleotide sequence that is the complement of a nucleotide sequence of (a)-(d).
 6. A gene chip comprising a nucleic acid molecule of claim
 5. 7. A transgenic non-human animal comprising a nucleic acid molecule of claim
 5. 8. A nucleic acid vector comprising a nucleic acid molecule of claim
 5. 9. A host cell containing the vector of claim
 8. 10. A method for producing any of the peptides of claim 1 comprising introducing a nucleotide sequence encoding any of the amino acid sequences in (a)-(d) into a host cell, and culturing the host cell under conditions in which the peptides are expressed from the nucleotide sequence.
 11. A method for producing any of the peptides of claim 2 comprising introducing a nucleotide sequence encoding any of the amino acid sequences in (a)-(d) into a host cell, and culturing the host cell under conditions in which the peptides are expressed from the nucleotide sequence.
 12. A method for detecting the presence of any of the peptides of claim 2 in a sample, said method comprising contacting said sample with a detection agent that specifically allows detection of the presence of the peptide in the sample and then detecting the presence of the peptide.
 13. A method for detecting the presence of a nucleic acid molecule of claim 5 in a sample, said method comprising contacting the sample with an oligonucleotide that hybridizes to said nucleic acid molecule under stringent conditions and determining whether the oligonucleotide binds to said nucleic acid molecule in the sample.
 14. A method for identifying a modulator of a peptide of claim 2, said method comprising contacting said peptide with an agent and determining if said agent has modulated the function or activity of said peptide.
 15. The method of claim 14, wherein said agent is administered to a host cell comprising an expression vector that expresses said peptide.
 16. A method for identifying an agent that binds to any of the peptides of claim 2, said method comprising contacting the peptide with an agent and assaying the contacted mixture to determine whether a complex is formed with the agent bound to the peptide.
 17. A pharmaceutical composition comprising an agent identified by the method of claim 16 and a pharmaceutically acceptable carrier therefor.
 18. A method for treating a disease or condition mediated by a human enzyme protein, said method comprising administering to a patient a pharmaceutically effective amount of an agent identified by the method of claim
 16. 19. A method for identifying a modulator of the expression of a peptide of claim 2, said method comprising contacting a cell expressing said peptide with an agent, and determining if said agent has modulated the expression of said peptide.
 20. An isolated human enzyme peptide having an amino acid sequence that shares at least 70% homology with an amino acid sequence shown in SEQ ID NO:2.
 21. A peptide according to claim 20 that shares at least 90 percent homology with an amino acid sequence shown in SEQ ID NO:2.
 22. An isolated nucleic acid molecule encoding a human enzyme peptide, said nucleic acid molecule sharing at least 80 percent homology with a nucleic acid molecule shown in SEQ ID NOS:1 or
 3. 23. A nucleic acid molecule according to claim 22 that shares at least 90 percent homology with a nucleic acid molecule shown in SEQ ID NOS:1 or
 3. 